BIX02188 (BIX-02188), an indolinone analog, is a novel, highly potent and selective MEK5 inhibitor that has the potential to treat neuropathic pain. MEK5 is inhibited with an IC50 of 4.3 nM, and ERK5 is also inhibited with an IC50 of 810 nM, but JNK2 and other closely related kinases are unaffected. By specifically inhibiting p-ERK5 expression in CSF-CN, BIX02188 can significantly reduce mechanical allodynia and thermal hypersensitivity brought on by CCI. This is accompanied by a decline in the phosphorylation of cAMP response-element binding protein (CREB) in CSF-CN.
Physicochemical Properties
| Molecular Formula | C25H24N4O2 |
| Molecular Weight | 412.48 |
| Exact Mass | 412.189 |
| Elemental Analysis | C, 72.80; H, 5.86; N, 13.58; O, 7.76 |
| CAS # | 334949-59-6 |
| Related CAS # | (E/Z)-BIX02188;1094614-84-2 |
| PubChem CID | 135398492 |
| Appearance | Light yellow to yellow solid powder |
| Density | 1.3±0.1 g/cm3 |
| Boiling Point | 615.8±55.0 °C at 760 mmHg |
| Flash Point | 326.2±31.5 °C |
| Vapour Pressure | 0.0±1.8 mmHg at 25°C |
| Index of Refraction | 1.688 |
| LogP | 2.36 |
| Hydrogen Bond Donor Count | 3 |
| Hydrogen Bond Acceptor Count | 4 |
| Rotatable Bond Count | 6 |
| Heavy Atom Count | 31 |
| Complexity | 645 |
| Defined Atom Stereocenter Count | 0 |
| SMILES | O=C(N)C1=CC(NC2=O)=C(/C2=C(C3=CC=CC=C3)/NC4=CC=CC(CN(C)C)=C4)C=C1 |
| InChi Key | WGPXKFOFEXJMBD-UHFFFAOYSA-N |
| InChi Code | InChI=1S/C25H24N4O2/c1-29(2)15-16-7-6-10-19(13-16)27-23(17-8-4-3-5-9-17)22-20-12-11-18(24(26)30)14-21(20)28-25(22)31/h3-14,28,31H,15H2,1-2H3,(H2,26,30) |
| Chemical Name | 3-[N-[3-[(dimethylamino)methyl]phenyl]-C-phenylcarbonimidoyl]-2-hydroxy-1H-indole-6-carboxamide |
| Synonyms | BIX-02188; BIX 02188; BIX02188 |
| 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) |
Biological Activity
| Targets | MEK5 (IC50 = 4.3 nM); ERK5 (IC50 = 810 nM); CSF1R (FMS) (IC50 = 280 nM); LCK (IC50 = 390 nM); KIT (IC50 = 550 nM); TGFβR1 (IC50 = 1.8 μM); ABL1 (IC50 = 2.1 μM); RPS6KA6 (RSK4) (IC50 = 3.2 μM); RPS6KA3 (RSK2) (IC50 = 4.1 μM); MAPK14 (p38 alpha) (IC50 = 3.9 μM); JAK3 (IC50 = 7.8 μM); SRC (IC50 = 8.9 μM) | ||
| ln Vitro | BIX02188 significantly inhibits ERK5 catalytic activity with an IC50 of 0.83 μM and blocks MEK5 catalytic activity with an IC50 of 4.3 nM. It exhibits no activity against the closely related kinases MEK1, MEK2, ERK1, p38α, JNK2, TGFβR1, EGFR, and STK16, with IC50 values of 1.8 μM for TGFβR1, 3.9 μM for p38α, and >6.3 μM for other kinases. Pretreatment with BIX02188 inhibits sorbitol-induced ERK5 phosphorylation in HeLa cells in a dose-dependent manner but has no inhibitory effect on ERK1/2, p38, or JNK1/2 MAPK phosphorylation. In HeLa or HEK293 cells, BIX02188 treatment given alone for 24 hours had no cytotoxic effects.[1] BIX02188 specifically inhibits BMK1 phosphorylation in bovine lung microvascular endothelial cells (BLMECs) stimulated with 300 M H2O2 in a dose-dependent manner, with an IC50 of 0.8 M. The fact that BIX02188 completely reverses the inhibitory effect on TNF-mediation and that JNK activation had a similar impact on BMK1 inhibition suggests that it inhibits TNF signaling by activating the MEK5-BMK1 signaling pathway.[2] | ||
| ln Vivo |
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| Enzyme Assay | Utilizing PKLight ATP Detection Reagent, kinase activity is measured for the MEK5 protein that was isolated from a baculovirus expression system. The assay is carried out in the presence of varying concentrations of BIX02188 using 15 nM GST-MEK5 and 0.75 μM ATP in an assay buffer made up of 25 mM Hepes, pH 7.5, 10 mM MgCl2, 50 mM KCl, 0.2% BSA, 0.01% CHAPS, 100 M Na3VO4, 0.5 mM DTT, and 1% DMSO. The kinase reaction mixture is incubated for 90 minutes at room temperature, and then 10 μL of an ATP detection reagent is added. For the purpose of determining the IC50 value, the relative light unit (RLU) signal is measured and converted to percent of control (POC) values. | ||
| Cell Assay | To stimulate the cells, sorbitol at a final concentration of 0.4 M is applied for 20 minutes at 37 °C after the cells have been serum-starved for 20 hours. 1.5 hours are spent adding BIX02188 before sorbitol is added. A 50 L RIPA buffer containing Halt protease and phosphate inhibitors is used to lyse the cells after cell harvesting for 5–10 minutes at 4 °C. The lysates are centrifuged for 10 minutes at 14,000 rpm, and then 50 μL of the lysate is mixed with 50 μl 2× sample buffer before boiling it for 4 minutes at 95 °C. Run on 10% Tris-glycine SDS-PAGE gels with nitrocellulose after a 20 µl sample. The proper antibodies are used for Western blotting. | ||
| Animal Protocol |
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| References |
[1]. Biochem Biophys Res Commun . 2008 Dec 5;377(1):120-5. [2]. Biochem Biophys Res Commun . 2008 May 23;370(1):159-63. |
Solubility Data
| Solubility (In Vitro) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 1.67 mg/mL (4.05 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 16.7 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 1.67 mg/mL (4.05 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 16.7 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. Solubility in Formulation 3: 30% PEG400+0.5% Tween80+5% propylene glycol: 30mg/mL (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.4244 mL | 12.1218 mL | 24.2436 mL | |
| 5 mM | 0.4849 mL | 2.4244 mL | 4.8487 mL | |
| 10 mM | 0.2424 mL | 1.2122 mL | 2.4244 mL |