Epoxomicin, a potent and selective proteasome inhibitor, exhibits<i>in vivo</i>antiinflammatory activity

Meng, Lihao; Mohan, Royce; Kwok, Benjamin H.; Elofsson, Mikael; Sin, Nancy L.; Crews, Craig M.

doi:10.1073/pnas.96.18.10403

Cited by 895 publications

(751 citation statements)

References 43 publications

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“…The natural product epoxomicin, a peptidyl epoxyketone, is the most selective proteasome inhibitor known due to its unique mechanism of inhibition (49,50). The final covalent adduct is a morpholino ring which contains not only the Thr hydroxyl, but also the N-terminal amino group (51).…”

Section: Discussionmentioning

confidence: 99%

Identification of the Proteasome Inhibitor MG262 as a Potent ATP-Dependent Inhibitor of the Salmonella enterica serovar Typhimurium Lon Protease

2006

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Lon is a homo-oligomeric ATP-dependent serine protease which functions in the degradation of damaged and certain regulatory proteins. The importance of Lon activity in bacterial pathogenicity has led to its emergence as a target in the development of novel antibiotics. As no potent inhibitors of Lon activity have been reported to date, we sought to identify an inhibitor which could serve as a lead compound in the development of a potent Lon-specific inhibitor. To determine whether a nucleotide-or peptide-based inhibitor would be more effective, we evaluated the steady-state kinetic parameters associated with both ATP and peptide hydrolysis by human and Salmonella enterica serovar Typhimurium Lon. Although the ATP hydrolysis activities of both homologs are kinetically indistinguishable, they display marked differences in peptide substrate specificity. This suggests that a peptide-based inhibitor could be developed which would target bacterial Lon, thereby decreasing side-effects due to cross-reactivity with human Lon. Using Salmonella enterica serovar Typhimurium Lon as a model, we evaluated the IC 50 values of a series of commercially available peptide-based inhibitors. Those inhibitors which behave as transition state analogs were the most useful in inhibiting Lon activity. The peptidyl boronate, MG262, was the most potent inhibitor tested (IC 50 = 122 ± 9 nM) and required binding, but not hydrolysis of ATP to initiate inhibition. We hope to use MG262 as a lead compound in the development of future Lon specific inhibitors.The number of pathogenic, antibiotic resistant bacteria increases each year, however the development of new antibiotics to treat them lags behind (1). Recent studies aimed at identifying proteins necessary for virulence have implicated the importance of Lon protease (2,3). Pathogenic Salmonella enterica are responsible for causing a range of human diseases from mild gastroenteritis (serovar Typhimurium and serovar Enteritidis) to typhoid fever (serovar Typhi). It has been demonstrated that Salmonella enteria serovar Typhimurium (S. Typhimurium) Lon protease activity is required for systemic infection in mice, a common study model for S. Typhi infection in humans (3). In fact, Lon-deficient S. Typhimurium, when administered as an oral vaccine to mice, conferred subsequent protection against infection by virulent S. Typhimurium (4). Taken together, these studies highlight Lon as an important target in the development of novel therapeutic agents.Lon, also known as the protease La, is a homo-oligomeric ATP-dependent serine protease, which functions in the degradation of damaged and certain short-lived regulatory proteins (5)(6)(7)(8)(9)(10)(11)(12)(13)(14). Homologs exist ubiquitously in nature, however they localize to the cytosol in prokaryotes and to the mitochondrial matrix in eukaryotes (8,15,16 (11,(18)(19)(20)(21)(22)(23).Lon protease is a member of the AAA + superfamily (ATPases Associated with different cellular Activities) along with other ATP-dependent proteases such as ClpXP, H...

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Section: Discussionmentioning

confidence: 99%

Identification of the Proteasome Inhibitor MG262 as a Potent ATP-Dependent Inhibitor of the Salmonella enterica serovar Typhimurium Lon Protease

2006

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“…(In this culture model, an increase of AβPP and Aβ is visible by immunoblots and Elisa within 24-48 hours after AβPP gene-transfer, and morphologic aspects of IBM appear approximately 14-21 days after the transfer.) Three days after transfer, experimental and control (non-AβPP-overexpressing) cultures were treated with 1μM epoxomicin (Biomol Research Laboratories, Plymouth Meeting, PA), an irreversible proteasome inhibitor [22]. 24 hours thereafter, the cultures were processed for light-microscopic immunocytochemistry, immunoblotting, and combined immunoprecipitation/immunoblotting, as described [4][5][6][7]11,23,24].…”

Section: Cultured Human Muscle Fibers (Chmfs)mentioning

confidence: 99%

AβPP-overexpression and proteasome inhibition increase αB-crystallin in cultured human muscle: Relevance to inclusion-body myositis

Wójcik

Engel

McFerrin

et al. 2006

Neuromuscular Disorders

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Amyloid-β precursor protein (AβPP) and its fragment amyloid-β (Aβ) are increased in s-IBM muscle fibers and appear to play an important role in the pathogenic cascade. αB-crystallin (αBC) was shown immunohistochemically to be accumulated in s-IBM muscle fibers, but the stressor(s) influencing αBC accumulation was not identified. We now demonstrate, using our experimental IBM model based on genetic overexpression of AβPP into normal culture human muscle fibers, that: 1) AβPP overexpression increased αBC 3.7-fold (p= 0.025); 2) additional inhibition of proteasome with epoxomicin increased αBC 7-fold (p=0.002); and 3) αBC physically associated with AβPP and Aβ oligomers. We also show that in biopsied s-IBM muscle fibers, αBC was similarly increased 3-fold (p=0.025) and physically associated with AβPP and Aβ oligomers. We propose that increased AβPP is a stressor increasing αBC expression in s-IBM muscle fibers. Determining the consequences of αBC association with Aβ oligomers could have clinical therapeutic relevance.

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“…Verification that solely proteasome activity contributed to the measured proteolytic activity was achieved by running parallel assays containing the highly specific proteasome inhibitor epoxomicin (PeptaNova, 4381) (Meng et al, 1999;Götze et al, 2013). All assays were performed in 96-well plates in a total volume of 50 μL per assay, containing 40 μL of assay buffer (0.05 M Tris(hydroxymethyl)-aminomethane, 25 mM KCl, 10 mM NaCl, 1 mM MgCl 2 , and 2 mM adenosine triphosphate (ATP), pH 8.0 at 30°C) and 5 μL of sample extract.…”

Section: Measurements Of Proteasome Activitiesmentioning

confidence: 99%

The proteasomes of two marine decapod crustaceans, European lobster (Homarus gammarus) and Edible crab (Cancer pagurus), are differently impaired by heavy metals

Götze

Bose

Sokolova

et al. 2014

Comparative Biochemistry and Physiology Part C: Toxicology &

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The intracellular ubiquitin-proteasome system is a key regulator of cellular processes involved in the controlled degradation of short-living or malfunctioning proteins. Certain diseases and cellular dysfunctions are known to arise from the disruption of proteasome pathways. Trace metals are recognized stressors of the proteasome system in vertebrates and plants, but their effects on the proteasome of invertebrates are not well understood. Since marine invertebrates, and particularly benthic crustaceans, can be exposed to high metal levels, we studied the effects of in vitro exposure to Hg 2+ , Zn 2+ , Cu 2+ , and Cd 2+ on the activities of the proteasome from the claw muscles of lobsters (Homarus gammarus) and crabs (Cancer pagurus). The chymotrypsin like activity of the proteasome of these two species showed different sensitivity to metals. In lobsters the activity was significantly inhibited by all metals to a similar extent. In crabs the activities were severely suppressed only by Hg 2+ and Cu 2+ while Zn 2+ had only a moderate effect and Cd 2+ caused almost no inhibition of the crab proteasome. This indicates that the proteasomes of both species possess structural characteristics that determine different susceptibility to metals. Consequently, the proteasome-mediated protein degradation in crab C. pagurus may be less affected by metal pollution than that of the lobster H. gammarus.

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Epoxomicin, a potent and selective proteasome inhibitor, exhibitsin vivoantiinflammatory activity

Cited by 895 publications

References 43 publications

Identification of the Proteasome Inhibitor MG262 as a Potent ATP-Dependent Inhibitor of the Salmonella enterica serovar Typhimurium Lon Protease

Identification of the Proteasome Inhibitor MG262 as a Potent ATP-Dependent Inhibitor of the Salmonella enterica serovar Typhimurium Lon Protease

AβPP-overexpression and proteasome inhibition increase αB-crystallin in cultured human muscle: Relevance to inclusion-body myositis

The proteasomes of two marine decapod crustaceans, European lobster (Homarus gammarus) and Edible crab (Cancer pagurus), are differently impaired by heavy metals

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