Ulixertinib (BVD-523)

Catalog No.S7854 Batch:S785406

Technical Data

Formula

C₂₁H₂₂Cl₂N₄O₂

Molecular Weight

433.33

CAS No.

869886-67-9

Solubility (25°C)*

In vitro

DMSO

87 mg/mL (200.77 mM)

Ethanol

22 mg/mL (50.76 mM)

Water

Insoluble

In vivo (Add solvents to the product individually and in order)

Clear solution

5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O

4.3mg/ml

Taking the 1 mL working solution as an example, add 50 μL 86 mg/ml clarified DMSO stock solution to 400 μL PEG300, mix evenly to clarify it; add 50 μL Tween80 to the above system, mix evenly to clarify it; then continue to add 500 μL ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.

* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

Preparing Stock Solutions

Biological Activity

Description

Ulixertinib (BVD-523, VRT752271) is a potent and reversible ERK1/ERK2 inhibitor with IC50 of <0.3 nM for ERK2. Phase 1.

Targets

ERK1 ^[1]	ERK2 ^[1]
	<0.3 nM

In vitro

In an A375 melanoma cell line containing a b-RAFV600E mutation, Ulixertinib reduces the levels of phosphorylated ERK2 (pERK) and of the phosphorylation of the downstream kinase RSK (pRSK) with IC50 of 4.1/0.14 μM, respectively. Ulixertinib also inhibits A375 cell proliferation with IC50 of 180 nM. ^[1]

Protocol (from reference)

Kinase Assay:^{^[1]}	ERK2 Rapidfire Mass Spectrometry Inhibition of Catalysis Assay MEK U911-activated ERK2 protein is expressed and purified in-house. Enzyme and substrate solutions are made up in assay buffer consisting of 50 mM Tris (pH 7.5), 10 mM MgCl₂, 0.1 mM EGTA, 10 mM DTT and 0.01% (v/v) CHAPS. 1.2 nM ERK2 protein is prepared in assay buffer and 10 µL is dispensed into each well of a polypropylene, 384-well plate containing test and reference control compounds. The compound plates had previously been dosed with a 12 point range from 100 µM down to 0.1 nM in order to calculate compound IC50s, with a total DMSO concentration in the assay of 1%. Following a 20 minute pre-incubation of enzyme and compound at room temperature, 10 µL of substrate solution is added consisting of 16 µM Erktide (IPTTPITTTYFFFK) and 120 µM ATP (measured Km) in assay buffer. The reaction is allowed to progress for 20 minutes at room temperature before being quenched by the addition of 80 µl 1% (v/v) formic acid. The assay plates are then run on the RapidFire Mass Spectrometry platform to measure substrate (unphosphorylated Erktide) and product (phosphorylated Erktide) levels.
Cell Assay:^{^[1]}	Cell lines A375 cells Concentrations ~30 μM Incubation Time 72 h Method A375 cells are cultured in cell media composed of DMEM, 10% (v/v) Foetal Calf Serum and 1% (v/v) L-Glutamine. After harvesting, cells are dispensed into black, 384-well Costar plates to give 200 cells per well in a total volume of 40 µL cell media, and are incubated overnight at 37°C, 90% relative humidity and 5% CO₂ in a rotating incubator. Test compounds and reference controls are dosed directly into the cell plates, into the inner 308 wells, using a Labcyte Echo 555 acoustic dispenser. The cells are dosed over a 12 point range from 30 µM down to 0.03 nM in order to calculate compound IC50s, with a total DMSO concentration in the assay of 0.3%. The cell plates are then incubated for 72 hours at 37°C. Cells were fixed and stained by the addition of 20 µL 12% formaldehyde in PBS/A (4% final concentration) and 1:2000 dilution of Hoechst 33342, with a 30 minute room temperature incubation, and then washed with PBS/A. A cell count is performed on the stained cell plates using a Cellomics ArrayScanTM VTI imaging platform. A Day 0 cell plate is also fixed, stained and read to generate a cell count baseline for determining compound cytotoxic effects as well as anti-proliferative effects.

Kinase Assay:^{^[1]}

ERK2 Rapidfire Mass Spectrometry Inhibition of Catalysis Assay
MEK U911-activated ERK2 protein is expressed and purified in-house. Enzyme and substrate solutions are made up in assay buffer consisting of 50 mM Tris (pH 7.5), 10 mM MgCl₂, 0.1 mM EGTA, 10 mM DTT and 0.01% (v/v) CHAPS. 1.2 nM ERK2 protein is prepared in assay buffer and 10 µL is dispensed into each well of a polypropylene, 384-well plate containing test and reference control compounds. The compound plates had previously been dosed with a 12 point range from 100 µM down to 0.1 nM in order to calculate compound IC50s, with a total DMSO concentration in the assay of 1%. Following a 20 minute pre-incubation of enzyme and compound at room temperature, 10 µL of substrate solution is added consisting of 16 µM Erktide (IPTTPITTTYFFFK) and 120 µM ATP (measured Km) in assay buffer. The reaction is allowed to progress for 20 minutes at room temperature before being quenched by the addition of 80 µl 1% (v/v) formic acid. The assay plates are then run on the RapidFire Mass Spectrometry platform to measure substrate (unphosphorylated Erktide) and product (phosphorylated Erktide) levels.

Cell Assay:^{^[1]}

Cell lines
A375 cells
Concentrations
~30 μM
Incubation Time
72 h
Method
A375 cells are cultured in cell media composed of DMEM, 10% (v/v) Foetal Calf Serum and 1% (v/v) L-Glutamine. After harvesting, cells are dispensed into black, 384-well Costar plates to give 200 cells per well in a total volume of 40 µL cell media, and are incubated overnight at 37°C, 90% relative humidity and 5% CO₂ in a rotating incubator. Test compounds and reference controls are dosed directly into the cell plates, into the inner 308 wells, using a Labcyte Echo 555 acoustic dispenser. The cells are dosed over a 12 point range from 30 µM down to 0.03 nM in order to calculate compound IC50s, with a total DMSO concentration in the assay of 0.3%. The cell plates are then incubated for 72 hours at 37°C. Cells were fixed and stained by the addition of 20 µL 12% formaldehyde in PBS/A (4% final concentration) and 1:2000 dilution of Hoechst 33342, with a 30 minute room temperature incubation, and then washed with PBS/A. A cell count is performed on the stained cell plates using a Cellomics ArrayScanTM VTI imaging platform. A Day 0 cell plate is also fixed, stained and read to generate a cell count baseline for determining compound cytotoxic effects as well as anti-proliferative effects.

References

[1] Ward RA, et al. J Med Chem. 2015, 58(11), 4790-4801.

Customer Product Validation

: Data from [Data independently produced by , , Blood, 2018, 131(18):2047-2059]

: Data from [Data independently produced by , , Haematologica, 2018, doi:10.3324/haematol.2018.196931]

: Data from [Data independently produced by , , Int J Clin Exp Med, 2016, 9(6):10955-10962.]

Selleck's Ulixertinib (BVD-523) has been cited by 69 publications

Tumour-selective activity of RAS-GTP inhibition in pancreatic cancer [ Nature, 2024, 629(8013):927-936]	PubMed: 38588697
EHMT2 promotes tumorigenesis in GNAQ/11-mutant uveal melanoma via ARHGAP29-mediated RhoA pathway [ Acta Pharm Sin B, 2024, 14(3):1187-1203]	PubMed: 38486999
Inhibition of oxidative stress-induced epithelial-mesenchymal transition in retinal pigment epithelial cells of age-related macular degeneration model by suppressing ERK activation [ J Adv Res, 2024, 60:141-157]	PubMed: 37328058
The mTOR pathway controls phosphorylation of BRAF at T401 [ Cell Commun Signal, 2024, 22(1):428]	PubMed: 39223665
Antitumor activity of extracellular signal-regulated kinases 1/2 inhibitor BVD-523 (ulixertinib) on thyroid cancer cells [ J Cancer Res Ther, 2024, 20(2):570-577]	PubMed: 38687926
ERK mediates interferon gamma-induced melanoma cell death [ Mol Cancer, 2023, 10.1186/s12943-023-01868-x]	PubMed: 37803324
Kinase-Modulated Bioluminescent Indicators Enable Noninvasive Imaging of Drug Activity in the Brain [ ACS Cent Sci, 2023, 9(4):719-732]	PubMed: 37122464
Kinase-Modulated Bioluminescent Indicators Enable Noninvasive Imaging of Drug Activity in the Brain [ ACS Cent Sci, 2023, 9(4):719-732]	PubMed: 37122464
Loss of NF1 in Melanoma Confers Sensitivity to SYK Kinase Inhibition [ Cancer Res, 2023, 83(2):316-331]	PubMed: 36409827
Impaired PPARγ activation by cadmium exacerbates infection-induced lung injury [ JCI Insight, 2023, 8(9)e166608]	PubMed: 36928191

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