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LED209

Alias: LED 209; LED-209; LED209
Cat No.:V12076 Purity: ≥98%
LED209 is a novel non-toxic inhibitor of the binding signals to QseC, preventing its autophosphorylation and consequently inhibiting QseC-mediated activation of virulence gene expression.
LED209
LED209 Chemical Structure CAS No.: 245342-14-7
Product category: Bacterial
This product is for research use only, not for human use. We do not sell to patients.
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Purity & Quality Control Documentation

Purity: ≥98%

Product Description
LED209 is a prodrug with strong selective action against the bacterial receptor QseC.It is a powerful small molecule inhibitor of QseC. Target: In vitro and in rodents, antibacterial LED209 exhibits no toxicity and has favorable pharmacokinetics. This is a novel antivirulence strategy that has been shown to be effective against a variety of Gram-negative pathogens that infect mammals over a wide range of conditions. Using a prodrug scaffold to deliver a warhead that allosterically modifies QseC and inhibits virulence in multiple Gram-negative pathogens, the LED209 QseC inhibitor has a novel mode of action.
Biological Activity I Assay Protocols (From Reference)
These protocols are for reference only. InvivoChem does not independently validate these methods.
Physicochemical Properties
Molecular Formula
C19H17N3O2S2
Molecular Weight
383.484
Exact Mass
383.076
Elemental Analysis
C, 59.51; H, 4.47; N, 10.96; O, 8.34; S, 16.72
CAS #
245342-14-7
Related CAS #
245342-14-7;
PubChem CID
3421033
Appearance
Solid powder
Density
1.4±0.1 g/cm3
Boiling Point
549.4±60.0 °C at 760 mmHg
Flash Point
286.1±32.9 °C
Vapour Pressure
0.0±1.5 mmHg at 25°C
Index of Refraction
1.741
LogP
3.4
Hydrogen Bond Donor Count
3
Hydrogen Bond Acceptor Count
4
Rotatable Bond Count
5
Heavy Atom Count
26
Complexity
542
Defined Atom Stereocenter Count
0
SMILES
O=S(C1=CC=C(NC(NC2=CC=CC=C2)=S)C=C1)(NC3=CC=CC=C3)=O
InChi Key
HNDRSTUKPCLQLT-UHFFFAOYSA-N
InChi Code
InChI=1S/C19H17N3O2S2/c23-26(24,22-17-9-5-2-6-10-17)18-13-11-16(12-14-18)21-19(25)20-15-7-3-1-4-8-15/h1-14,22H,(H2,20,21,25)
Chemical Name
1-Phenyl-3-[4-(phenylsulfamoyl)phenyl]thiourea
Synonyms
LED 209; LED-209; LED209
HS Tariff Code
2934.99.9001
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)
Solubility Data
Solubility (In Vitro)
DMSO : ~250 mg/mL (~651.91 mM)
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
(e.g. IP/IV/IM/SC)
Injection Formulation 1: DMSO : Tween 80: Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution 50 μL Tween 80 850 μL Saline)
*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 : PEG300Tween 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).
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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)]
*Preparation of 20% SBE-β-CD in Saline (4°C,1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO 100 μLPEG300 200 μL castor oil 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10: 10 : 80 (i.e. 100 μL Ethanol 100 μL Cremophor 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH 400 μLPEG300 50 μL Tween 80 450 μL 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).
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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders


Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

 (Please use freshly prepared in vivo formulations for optimal results.)
Preparing Stock Solutions 1 mg 5 mg 10 mg
1 mM 2.6077 mL 13.0385 mL 26.0770 mL
5 mM 0.5215 mL 2.6077 mL 5.2154 mL
10 mM 0.2608 mL 1.3038 mL 2.6077 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.

Calculator

Molarity Calculator allows you to calculate the mass, volume, and/or concentration required for a solution, as detailed below:

  • Calculate the Mass of a compound required to prepare a solution of known volume and concentration
  • Calculate the Volume of solution required to dissolve a compound of known mass to a desired concentration
  • Calculate the Concentration of a solution resulting from a known mass of compound in a specific volume
An example of molarity calculation using the molarity calculator is shown below:
What is the mass of compound required to make a 10 mM stock solution in 5 ml of DMSO given that the molecular weight of the compound is 350.26 g/mol?
  • Enter 350.26 in the Molecular Weight (MW) box
  • Enter 10 in the Concentration box and choose the correct unit (mM)
  • Enter 5 in the Volume box and choose the correct unit (mL)
  • Click the “Calculate” button
  • The answer of 17.513 mg appears in the Mass box. In a similar way, you may calculate the volume and concentration.

Dilution Calculator allows you to calculate how to dilute a stock solution of known concentrations. For example, you may Enter C1, C2 & V2 to calculate V1, as detailed below:

What volume of a given 10 mM stock solution is required to make 25 ml of a 25 μM solution?
Using the equation C1V1 = C2V2, where C1=10 mM, C2=25 μM, V2=25 ml and V1 is the unknown:
  • Enter 10 into the Concentration (Start) box and choose the correct unit (mM)
  • Enter 25 into the Concentration (End) box and select the correct unit (mM)
  • Enter 25 into the Volume (End) box and choose the correct unit (mL)
  • Click the “Calculate” button
  • The answer of 62.5 μL (0.1 ml) appears in the Volume (Start) box
g/mol

Molecular Weight Calculator allows you to calculate the molar mass and elemental composition of a compound, as detailed below:

Note: Chemical formula is case sensitive: C12H18N3O4  c12h18n3o4
Instructions to calculate molar mass (molecular weight) of a chemical compound:
  • To calculate molar mass of a chemical compound, please enter the chemical/molecular formula and click the “Calculate’ button.
Definitions of molecular mass, molecular weight, molar mass and molar weight:
  • Molecular mass (or molecular weight) is the mass of one molecule of a substance and is expressed in the unified atomic mass units (u). (1 u is equal to 1/12 the mass of one atom of carbon-12)
  • Molar mass (molar weight) is the mass of one mole of a substance and is expressed in g/mol.
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Reconstitution Calculator allows you to calculate the volume of solvent required to reconstitute your vial.

  • Enter the mass of the reagent and the desired reconstitution concentration as well as the correct units
  • Click the “Calculate” button
  • The answer appears in the Volume (to add to vial) box
In vivo Formulation Calculator (Clear solution)
Step 1: Enter information below (Recommended: An additional animal to make allowance for loss during the experiment)
Step 2: Enter in vivo formulation (This is only a calculator, not the exact formulation for a specific product. Please contact us first if there is no in vivo formulation in the solubility section.)
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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.

Biological Data
  • LED209 targets the histidine sensor kinase QseC to attenuate virulence gene expression. (A, B) LED209 signals through the bacterial receptor QseC but not through the bacterial receptor QseE. Wild-type (WT) EHEC and the ΔqseC (A) and ΔqseE (B) isogenic mutants were grown in vitro in the presence (5 nM) or absence (DMSO only) of LED209 to late logarithmic phase. Expression of ler, the master transcriptional regulator of the LEE pathogenicity island in EHEC, of tir, a key LEE-encoded virulence gene, and of stx2a, a subunit of Shiga toxin, was measured by qRT-PCR. Expression is relative to that of the WT grown in the absence of LED209 (error bars, standard deviation [SD]; **, P < 0.01; ***, P < 0.001; NS, not significant). (C) QseC homologues are present in more than 25 plant and animal pathogens.[1].QseC inhibitors as an antivirulence approach for Gram-negative pathogens. MBio. 2014 Nov 11;5(6):e02165.
  • LED209 acts as a prodrug, cleaving an aniline group to expose its active component. (A) Upon interaction with QseC, LED209 breaks into the active component of LED209, OM188, and an aniline group. Schematic depicts the breakdown of LED209. OM188 contains an isothiocyanate (S=C=N) that is reactive to lysine and cysteine. (B) Treatment of EHEC with OM188 in vitro (by adding DMSO to the no-drug control and OM188 in DMSO at the beginning of the experiment) decreases expression of virulence genes. WT EHEC was grown in vitro in the presence (500 nM) or absence (DMSO only) of OM188 to late logarithmic phase. Expression of ler, eae, and stx2a was measured by qRT-PCR. Expression is relative to that of the WT grown in the absence of OM188 (error bars, SD; **, P < 0.01; *, P < 0.05). (C) Fluorescent actin staining assay of HeLa cells infected with EHEC grown in the absence (DMSO, no drug) or presence (5 nM) of OM188 or LED209, stained with fluorescein isothiocyanate-phalloidin (actin, green) and propidium iodide (bacterial and HeLa DNA, red). Original magnification, ×63. (D) Quantification of fluorescent actin staining assay of the percentage of HeLa cells infected, as defined by pedestal formation by EHEC (n = 100 cells). (E) Intramacrophage replication of S. enterica Typhimurium in the presence and absence of OM188 (error bars, SD; ***, P < 0.001; **, P < 0.01; *, P < 0.05).[1].QseC inhibitors as an antivirulence approach for Gram-negative pathogens. MBio. 2014 Nov 11;5(6):e02165.
  • Treatment of LED209 protects against S. Typhimurium and F. tularensis. (A, B) Administration of LED209 protects BALB/c mice from S. Typhimurium infection. Mice were administered a single dose of LED209 (20 mg/kg) at 3 h before (A) or multiple doses of LED209 (20 mg/kg) at 3 h before, at the time of, and 3, 6, 9, and 12 h after (B) intraperitoneal injection of a lethal dose of S. Typhimurium (106 CFU). Graphs indicate survival to infection (n = 10). (C) Treatment of S. Typhimurium with 50 nM LED209 in vitro decreased the expression of sifA, an effector required for vacuolar replication. qRT-PCR of sifA in S. Typhimurium grown in N-minimal medium to late logarithmic phase. Expression is relative to S. Typhimurium grown in the absence of drug (error bars, SD; ***, P < 0.001). (D) J774 murine macrophages were infected with opsonized S. Typhimurium in the presence (5 nM) or absence (DMSO only) of LED209. Graph indicates intracellular survival within macrophages. (E, F) Administration of LED209 postinfection protects C3H/HeN mice from F. tularensis infection. Mice were administered a single dose of LED209 (20 mg/kg) at 1, 3, 6, 9, or 24 h prior to infection and then intranasally infected with 30 CFU of F. tularensis strain SCHU S4 (E) or a single dose at 1, 3, 6, 9, or 24 h postinfection with 30 CFU of F. tularensis (F). Graphs indicate survival to infection (n = 10).[1].QseC inhibitors as an antivirulence approach for Gram-negative pathogens. MBio. 2014 Nov 11;5(6):e02165.
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