GW4869

[Description]:

GW4869 is a noncompetitive neutral sphingomyelinase inhibitor with an IC50 of 1 μM.

[Related Catalog]:

Signaling Pathways >> Others >> Others

Research Areas >> Cardiovascular Disease

Research Areas >> Inflammation/Immunology

[Target]

IC50: 1 μM (neutral sphingomyelinase)[1]

[In Vitro]

GW4869 (10 μM) partially inhibits TNF-induced sphingomyelin (SM) hydrolysis, and 20 μM of the compound is protected completely from the loss of SM. The addition of 10-20 μM GW4869 completely inhibits the initial accumulation of ceramide, whereas this effect is partially lost at later time points (24 h). The action of GW4869 occurs downstream of the drop in glutathione. GW4869 is able, in a dose-dependent manner, to significantly protect from cell death. These protective effects are accompanied by significant inhibition of cytochrome c release from mitochondria and caspase 9 activation, therefore localizing N-SMase activation upstream of mitochondrial dysfunction[1].

[In Vivo]

Pre-treatment with GW4869 significantly impairs release of both exosomes and pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) in RAW264.7 macrophages. At 12 h after LPS treatment or CLP surgery, WT mice pretreated with GW4869 displays lower amounts of exosomes and pro-inflammatory cytokines in the serum than control PBS-injected mice. Accordingly, GW4869 treatment diminishes the sepsis-induced cardiac inflammation, attenuates myocardial depression and prolongs survival[2].

[Kinase Assay]

B. cereu sphosphatidylcholine-PLC is incubated in the presence or absence of 10 μM GW4869 in a reaction mixture containing 100 mM Tris, pH 7.2, 25% glycerol, 20 mM p-nitrophenyl/phosphorylcholine, and production of p-nitrophenol is quantified spectrophotometrically at 410 nm. Protein phosphatase 2A from bovine kidney is incubated in the presence or absence of GW4869 in buffer containing 50 mM Tris, pH 7.4, 1 mM dithiothreitol, 100 μM MnCl2, and 20% glycerol, and phosphatase activity is measured[1].

[Cell Assay]

Cells are treated with GW4869 for 30 min and then TNF is added in 10 μL/well. At the indicated time points, 25 μL of MTT stock solution are added to each well and incubated at 37 °C in 5% CO2 for 3 h. The cell viability is using the MTT assay[1].

[Animal admin]

Mice: The mice are randomly assigned to four groups: PBS, GW4869, PBS+LPS and GW4869+LPS (n=5 per group). GW4869 is intraperitoneally (i.p.) injected at one dose of 2.5μg/g. Mice in the PBS+LPS group are pre-injected i.p. with PBS 1 h prior to an i.p. injection of LPS (25 μg/g). Mice in the group of GW4869+LPS are pre-injected i.p. with GW4869 (2.5μg/g) for 1 h, followed by an i.p. injection of LPS (25 μg/g, 100μl). Mice receive injections of PBS to a comparable volume (100μl) as controls. The survival rate of the PBS+LPS and GW4869+LPS groups are monitored every 6 h for a 36 h period[2].

[References]

[1]. Luberto C, et al. Inhibition of tumor necrosis factor-induced cell death in MCF7 by a novel inhibitor of neutralsphingomyelinase. J Biol Chem. 2002 Oct 25;277(43):41128-39.

[2]. Essandoh K, et al. Blockade of exosome generation with GW4869 dampens the sepsis-induced inflammation and cardiac dysfunction. Biochim Biophys Acta. 2015 Nov;1852(11):2362-71.

[3]. Nakamura H, et al. Sphingomyelin Regulates the Activity of Secretory Phospholipase A2 in the Plasma Membrane. J Cell Biochem. 2015 Sep;116(9):1898-907.

[Related Small Molecules]

[ Molecular Formula ]:
C₃₀H₃₀Cl₂N₆O₂

[ Molecular Weight ]:
577.50

[ PSA ]:
127.21000

[ LogP ]:
4.57730

[ Storage condition ]:
-20℃

[ Personal Protective Equipment ]:
Eyeshields;Gloves;type N95 (US);type P1 (EN143) respirator filter

[ RIDADR ]:
NONH for all modes of transport

Glutaminase-containing microvesicles from HIV-1-infected macrophages and immune-activated microglia induce neurotoxicity.

Molecular Neurodegeneration 10 , 61, (2015)

HIV-1-infected and/or immune-activated microglia and macrophages are pivotal in the pathogenesis of HIV-1-associated neurocognitive disorders (HAND). Glutaminase, a metabolic enzyme that facilitates g...

Regulation of SREBPs by Sphingomyelin in Adipocytes via a Caveolin and Ras-ERK-MAPK-CREB Signaling Pathway.

PLoS ONE 10 , e0133181, (2015)

Sterol response element binding protein (SREBP) is a key transcription factor in insulin and glucose metabolism. We previously demonstrated that elevated levels of membrane sphingomyelin (SM) were rel...

Secretion of small/microRNAs including miR-638 into extracellular spaces by sphingomyelin phosphodiesterase 3.

Oncol. Rep. 33(1) , 67-73, (2014)

A recent study demonstrated that intracellular small/microRNAs are released from cells, and some of these extracellular RNAs are embedded in vesicles, such as ceramide-rich exosomes, on lipid-bilayer ...

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