IC50: 30 nM (FLAP)[3] IC50: 3 nM (Leukotriene biosynthesis in intact leukocytes) and 1.1 μM (Leukotriene biosynthesis in human whole blood)[2] PPARα[1]

In a culture of mouse primary keratinocytes, MK-886 sodium salt (0.5-2 μM; 15 hours) treatment lowers keratin-1 expression[1]. 10 μM MK-886 sodium salt can reduce Wy-14643 activation of PPARα by approximately 80% in monkey kidney fibroblast CV-1 cells, mouse keratinocyte 308 cells, and human lung adenocarcinoma A549 cells using a transient transfection technique. In the stable transfection system, MK-886 sodium salt also reduces PPARα activation by fatty acids [1]. Despite the expression of all PPAR isoforms in Jurkat cells, apoptosis induced by MK-886 sodium salt cannot be prevented by different PPARα and PPARγ agonists[1].

Male Sprague-Dawley rats treated with MK-886 sodium salt (L 663536; 5 mg/kg; oral administration) show a strong inhibition of antigen-induced dyspnea in inbred rats that have previously received methysergide treatment[2]. Using rat pleurisy (ED50, 0.2 mg/kg po), inflammatory paw (ED50, 0.8 mg/kg), and a model of leukotriene excretion in rat bile after antigen provocation, MK-886 sodium salt (L 663536) suppresses leukotriene production in vivo[2].

PPAR ligand binding assay: Binding of MK886 to PPARs was determined using the coactivator-dependent receptor ligand assay (CARLA). A construct containing the PPARα ligand binding domain cloned in frame with glutathione S-transferase (GST) was procured from Walter Wahli. The GST–PPAR ligand binding domain fusion protein was expressed in Escherichia coli BL2 DE3 (pLysS). Bacterial pellets containing the fusion protein were resuspended in 10 ml of lysis buffer [PBS containing 1% (v}v) Triton X-100 and 0.5 mM PMSF] and lysed by repeated freeze–thawing. DNA and insoluble matter were removed by centrifugation at 4500 g. The fusion protein was purified using GSH–Sepharose beads at 4 °C, washed three times in lysis buffer, and equilibrated in 20 mM Tris}HCl (pH 8.0), 100 mM NaCl, 1 mM EDTA, 0.5% Nonidet P-40 and 1 mM dithiothreitol (DTT) supplemented with 1% (w}v) dry milk. The amount of protein used per reaction was 1–3 µg. The beads were incubated with different concentrations of MK886 and $&S-radiolabelled steroid receptor coactivator-1 (SRC-1) prepared with a pSG5 plasmid construct that can express SRC-1 (procured from Walter Wahli) in Šitro using a coupled transcription}translation rabbit reticulocyte lysate system. Labelling was achieved by incubating for 1 h in the presence of [$&S]methionine, and the beads were recovered by centrifugation. The beads were then washed and analysed for interaction with SRC-1 using SDS}PAGE. Coomassie Brilliant Blue staining of GST–ligand binding domain fusions allowed standardization between different reactions. The SRC-1 protein complex was visualized by autoradiography. Relative band densities were determined using a scanned image and UN-SCAN-IT software[1].

Western Blot Analysis[1]
Cell Types: Primary Keratinocytes
Tested Concentrations: 0.5 µM, 1 µM or 2 µM
Incubation Duration: 15 hrs (hours)
Experimental Results: diminished keratin-1 expression.

Animal/Disease Models: Male SD (SD (Sprague-Dawley)) rat (300-400 g), antigen-induced dyspnea [1]
Doses: 5 mg/kg
Route of Administration: Oral
Experimental Results:Inhibition of antigen-induced dyspnea.

[1]. Inhibition of peroxisome-proliferator-activated receptor (PPAR)alpha by MK886. Biochem J. 2001 Jun 15;356(Pt 3):899-906.
[2]. L-663,536 (MK-886) (3-[1-(4-chlorobenzyl)-3-t-butyl-thio-5-isopropylindol-2-yl]-2,2 - dimethylpropanoic acid), a novel, orally active leukotriene biosynthesis inhibitor. Can J Physiol Pharmacol. 1989 May;67(5):456-64.
[3]. Mancini JA, et al. 5-Lipoxygenase-activating protein is the target of a novel hybrid of two classes of leukotriene biosynthesis inhibitors. Mol Pharmacol. 1992 Feb;41(2):267-72.

3-[3-(tert-butylsulfanyl)-1-(4-chlorobenzyl)-5-(propan-2-yl)-1H-indol-2-yl]-2,2-dimethylpropanoic acid is a member of the class of indoles that is 1H-indole substituted by a isopropyl group at position 5, a tert-butylsulfanediyl group at position 3, a 4-chlorobenzyl group at position 1 and a 2-carboxy-2-methylpropyl group at position 2. It acts as an inhibitor of arachidonate 5-lipoxygenase. It has a role as an EC 1.13.11.34 (arachidonate 5-lipoxygenase) inhibitor, an antineoplastic agent and a leukotriene antagonist. It is an aryl sulfide, a member of indoles, a monocarboxylic acid and a member of monochlorobenzenes.
MK-886 is an experimental inhibitor of leukotriene synthesis.

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Although MK886 was originally identified as an inhibitor of 5-lipoxygenase activating protein (FLAP), recent data demonstrate that this activity does not underlie its ability to induce apoptosis [Datta, Biswal and Kehrer (1999) Biochem. J. 340, 371--375]. Since FLAP is a fatty-acid binding protein, it is conceivable that MK886 may affect other such proteins. A family of nuclear receptors that are activated by fatty acids and their metabolites, the peroxisome-proliferator-activated receptors (PPARs), have been implicated in apoptosis and may represent a target for MK886. The ability of MK886 to inhibit PPAR-alpha, -beta and -gamma activity was assessed using reporter assay systems (peroxisome-proliferator response element--luciferase). Using a transient transfection system in monkey kidney fibroblast CV-1 cells, mouse keratinocyte 308 cells and human lung adenocarcinoma A549 cells, 10--20 microM MK886 inhibited Wy14,643 activation of PPAR alpha by approximately 80%. Similar inhibition of PPAR alpha by MK886 was observed with a stable transfection reporter system in CV-1 cells. Only minimal inhibitory effects were seen on PPAR beta and PPAR gamma. MK886 inhibited PPAR alpha by a non-competitive mechanism as shown by its effects on the binding of arachidonic acid to PPAR alpha protein, and a dose-response study using a transient transfection reporter assay in COS-1 cells. An assay assessing PPAR ligand-receptor interactions showed that MK886 prevents the conformational change necessary for active-complex formation. The expression of keratin-1, a protein encoded by a PPAR alpha-responsive gene, was reduced by MK886 in a culture of mouse primary keratinocytes, suggesting that PPAR inhibition has functional consequences in normal cells. Although Jurkat cells express all PPAR isoforms, various PPAR alpha and PPAR gamma agonists were unable to prevent MK886-induced apoptosis. This is consistent with MK886 functioning as a non-competitive inhibitor of PPAR alpha, but may also indicate that PPAR alpha is not directly involved in MK886-induced apoptosis. Although numerous PPAR activators have been identified, the results show that MK886 can inhibit PPAR alpha, making it the first compound identified to have such an effect.[1]

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L-663,536 (3-[1-(4-chlorobenzyl)-3-t-butyl-thio-5-isopropylindol-2-yl]-2, 2-dimethylpropanoic acid) is a potent inhibitor of leukotriene (LT) biosynthesis in intact human polymorphonuclear leukocytes (PMN) (IC50, 2.5 nM). Similarly, L-663,536 inhibited A23187-induced LTB4 formation by rat peripheral blood and elicited PMN. At concentrations where inhibition of leukotriene biosynthesis occurred in human whole blood (1.1 microM), no effect was seen on cyclooxygenase or 12-lipoxygenase, an effect also observed in washed human platelets. The compound had no effect on rat or porcine 5-lipoxygenase indicating that L-663,536 is not a direct 5-lipoxygenase inhibitor. When administered in vivo L-663,536 was a potent inhibitor of antigen-induced dyspnea in inbred rats pretreated with methysergide (ED50, 0.036 mg/kg p.o.) and of Ascaris-induced bronchoconstriction in squirrel monkeys (1 mg/kg p.o.). The compound inhibited leukotriene biosynthesis in vivo in a rat pleurisy model (ED50, 0.2 mg/kg p.o.), an inflamed rat paw model (ED50, 0.8 mg/kg), a model of leukotriene excretion in rat bile following antigen provocation, and a model in the guinea-pig ear where leukotriene synthesis was induced by topical challenge with ionophore A23187 (ED50, 2.5 mg/kg p.o. and 0.6 micrograms topically). The results indicate that L-663,536 is a potent inhibitor of leukotriene biosynthesis both in vitro and in vivo indicating that the compound is suitable for studying the role of leukotrienes in a variety of pathological situations.[2]

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An 18-kDa leukocyte membrane protein, termed 5-lipoxygenase-activating protein (FLAP), has recently been shown to be the target of two structurally distinct classes of leukotriene biosynthesis inhibitors. These classes of inhibitors are based on indole and quinoline structures and are represented by MK-886 and L-674,573, respectively. A novel class of hybrid structure based on the indole and quinoline classes of inhibitors, termed quindoles, has recently been developed. These compounds, exemplified by L-689,037, are potent inhibitors of leukotriene biosynthesis, both in vitro and in vivo. In the present study, we have developed and characterized a potent radioiodinated photoaffinity analogue of L-689,037, termed [125I]L-691,678. This compound was used in immunoprecipitation studies with FLAP antisera to show that the quindole series of leukotriene biosynthesis inhibitors interact directly with FLAP. In addition, we show that MK-886, L-674,573, and L-689,037 specifically compete, in a concentration-dependent manner, with both [125I]L-691,678 and [125I]L-669,083, a photoaffinity analogue of MK-886, for binding to FLAP. These results suggest that these three classes of leukotriene biosynthesis inhibitors share a common binding site on FLAP, providing further evidence that FLAP represents a suitable target for structurally diverse classes of leukotriene biosynthesis inhibitors.[3]

C27H33CLNNAO2S

494.06

493.181

C, 65.64; H, 6.73; Cl, 7.18; N, 2.84; Na, 4.65; O, 6.48; S, 6.49

118427-55-7

MK-886;118414-82-7

4519262

Typically exists as solid at room temperature

6.675

0

3

8

33

645

0

CC(C)C1=CC2=C(C=C1)N(CC3=CC=C(C=C3)Cl)C(=C2SC(C)(C)C)CC(C)(C)C(=O)[O-].[Na+]

CBNCIYNCWVGEKJ-UHFFFAOYSA-M

InChI=1S/C27H34ClNO2S.Na/c1-17(2)19-10-13-22-21(14-19)24(32-26(3,4)5)23(15-27(6,7)25(30)31)29(22)16-18-8-11-20(28)12-9-18;/h8-14,17H,15-16H2,1-7H3,(H,30,31);/q;+1/p-1

sodium;3-[3-tert-butylsulfanyl-1-[(4-chlorophenyl)methyl]-5-propan-2-ylindol-2-yl]-2,2-dimethylpropanoate

MK-886 sodium salt; 118427-55-7; MK886 sodium; MK-886 (sodium salt); PNR27O326B; sodium;3-[3-tert-butylsulfanyl-1-[(4-chlorophenyl)methyl]-5-propan-2-ylindol-2-yl]-2,2-dimethylpropanoate; CHEMBL416657; MK 886 sodium;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

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

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.

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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 : 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).

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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 : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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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

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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.

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 (Please use freshly prepared in vivo formulations for optimal results.)