| Size | Price | Stock | Qty |
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| 500mg |
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| 1g |
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| Other Sizes |
| Targets |
Hymecromone targets NSMase2 (neutral sphingomyelinase 2) as an activator (no IC50/Ki reported); downstream effectors include ceramide, PP2A, Akt (Ser473 phosphorylation inhibited), HAS2, calpain1/2, p53 (phosphorylated), caspase-3 (activated), and SIRT1 (downregulated). [1]
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| ln Vitro |
[1] Hymecromone (1 mM) decreased HA production in G26-24 cells from 3250±221 to 291±14 ng/mg cell protein (over 90% reduction) after 24 h. It reduced cell viability in a dose-dependent manner (0–1 mM) as measured by MTT assay, and induced apoptosis as shown by annexin V/PI flow cytometry; 2 mM MU also induced apoptosis. The NSMase2 inhibitor GW4869 (5 μM) partially reversed the loss of viability and apoptosis. MU treatment caused morphological changes (stretched and adherent with processes), which were partially reversed by GW4869. MU did not penetrate into cells (no fluorescence uptake after 3 h). MU activated NSMase2 (within 15 min) and elevated ceramide levels (total and individual species) in a time-dependent manner (0.25–24 h), as shown by HPTLC and LC/MS/MS; the ceramide/SM ratio increased dose- and time-dependently, and this was reversed by GW4869. MU increased PP2A expression and decreased phosphorylation of Akt (Ser473) at 24 h. MU reduced HAS2 expression at 0.5, 1, 3, and 24 h, which was partially reversed by GW4869. MU promoted translocation of NSMase2 to lipid rafts and increased its activity (up to 3 h) without affecting ASMase. Other NSMase2 activators (bacterial SMase, staurosporine, H2O2) and exogenous C2-ceramide also reduced HA synthesis and increased cell adherence. MU decreased migration (by ~24% at 0.5 mM and ~42% at 1 mM) and invasion (by ~30% and ~40%) of G26-24 cells. MU reduced calpain activity and increased pro-calpain1 and pro-calpain2 protein levels. MU increased p-P53, activated caspase-3 (activity and cleavage), and decreased SIRT1 expression. [1]
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| Enzyme Assay |
[1] NSMase2 and ASMase activities were determined using the fluorimetric substrate HMU-phosphorylcholine. Cell lysates (50 μg protein) were incubated with the substrate at pH 7.4 (for NSMase2, with 10 mM MgCl2, 100 mM Tris-HCl, 0.1% Triton X-100, and 5 mM DTT) or pH 4.5 (for ASMase, with 150 mM sodium acetate and 1 mM EDTA) ; the released HMU was measured fluorometrically (excitation/emission not specified, but using a microplate reader) and activity calculated from the slope of fluorescence vs. time, normalized to protein.
Calpain activity was measured using the cell-permeable substrate t-BOC-LM-CMAC (30 μM). Cells treated with 1 mM MU for 60 min were lysed, and the fluorescence of liberated CMAC was quantified (ex 351 nm, em 430 nm). Caspase-3 activity was assayed by incubating 25–30 μg of cell extract with DEVD-AFC substrate in HEPES buffer (pH 7.4, 2 mM DTT, 5 mM EDTA) at 37 °C for 3 h; fluorescence was measured at ex 400 nm, em 505 nm, and activity calculated from the slope normalized to protein. [1] |
| Cell Assay |
[1] HA production was measured by competitive ELISA: conditioned media from G26-24 cells (cultured for 48 h) were mixed with detector and added to HA-coated plates; colorimetric signal at 450 nm inversely correlated with HA amount, normalized to cell protein.
Cell viability was assessed by MTT assay: cells seeded in 24-well plates (1×10^5/cm²) were treated with MU (0–1 mM), then MTT (5 mg/mL) added for 2 h, followed by dissolution in 10% SDS-HCl and OD measurement at 595 nm. Apoptosis was analyzed by flow cytometry using Alexa Fluor 488 annexin V/Dead cell apoptosis kit; cells were stained and percentage of early and late apoptotic cells was calculated. MU uptake was measured by fluorescence (ex 360, em 460) in cell lysates and supernatants after sonication, and by confocal microscopy using DRAQ5 for nuclear staining; no uptake was detected after 3 h. Western blotting: cell lysates were separated by SDS-PAGE, transferred to membranes, and probed with antibodies against NSMase2, Akt, p-Akt (Ser473), HAS2, calpain1, calpain2, PP2A, caspase3, cleaved caspase3, p53, SIRT1, and β-actin; bands were quantified using densitometry. Lipid raft isolation: cells lysed in Triton X-100 at 4 °C, sucrose gradient ultracentrifugation, fractions collected and analyzed by Western blot. Lipid extraction and HPTLC: cells labeled with [³H]palmitate for 24 h, lipids extracted by chloroform-methanol-water, subjected to alkaline methanolysis, separated on HPTLC with chloroform/methanol/acetic acid/water, visualized with iodine, and quantified by scintillation counting to determine ceramide/SM ratio. LC/MS/MS: lipids extracted with 17:0-Cer as internal standard, analyzed by LC/MS/MS with ESI positive ion mode and MRM; standard curves (0–300 pmol) with R²>0.98 used for quantification of individual ceramide species. Migration and invasion assays were performed using CytoSelect 24-well kits per manufacturer's instructions; cells treated with MU (0.5 or 1 mM) for 24 h, then seeded in upper chambers and incubated; migrated/invaded cells were stained and quantified. [1] |
| Toxicity/Toxicokinetics |
Hymecromone is a HA synthesis inhibitor traditionally thought to act via UDP-glucuronic acid depletion, but this study reveals a novel mechanism: it activates NSMase2 on the plasma membrane, hydrolyzing sphingomyelin to ceramide, which then activates PP2A, dephosphorylates Akt, reduces HAS2 expression and HA synthesis; PP2A also inhibits calpain1/2, reducing migration and invasion; ceramide further activates p53 and caspase-3 while suppressing SIRT1, leading to apoptosis. This mechanism explains both anti-metastatic and pro-apoptotic effects of MU. The compound does not need to enter cells to initiate these signaling events, as it acts on the cell surface. This work suggests that ceramide-generating drugs (NSMase2 activators) may be useful for cancer therapy. [1]
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| References |
Biochim Biophys Acta.2016Feb;1861(2):78-90.
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| Molecular Formula |
C10H7NAO3
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|---|---|
| Molecular Weight |
198.15
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| Exact Mass |
198.029
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| CAS # |
5980-33-6
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| PubChem CID |
3364573
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| Appearance |
Typically exists as solid at room temperature
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| Density |
1.319g/cm3
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| Boiling Point |
377.4ºC at 760 mmHg
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| Melting Point |
90-92ºC(lit.)
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| Flash Point |
174.5ºC
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| Index of Refraction |
1.611
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| LogP |
2.245
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| Hydrogen Bond Donor Count |
0
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| Hydrogen Bond Acceptor Count |
3
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| Rotatable Bond Count |
0
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| Heavy Atom Count |
14
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| Complexity |
262
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| Defined Atom Stereocenter Count |
0
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| SMILES |
0
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| InChi Key |
JGMQHDNPUCPRQE-UHFFFAOYSA-M
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| InChi Code |
InChI=1S/C10H8O3.Na/c1-6-4-10(12)13-9-5-7(11)2-3-8(6)9;/h2-5,11H,1H3;/q;+1/p-1
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| Chemical Name |
sodium;4-methyl-2-oxochromen-7-olate
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| Synonyms |
Hymecromone sodium 4-Methylumbelliferone sodium
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 5.0467 mL | 25.2334 mL | 50.4668 mL | |
| 5 mM | 1.0093 mL | 5.0467 mL | 10.0934 mL | |
| 10 mM | 0.5047 mL | 2.5233 mL | 5.0467 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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