TG003

Catalog No.S7320

For research use only.

TG003 is a potent and ATP-competitive Cdc2-like kinase (Clk) inhibitor with IC50 of 20 nM, 200 nM, and 15 nM for Clk1, Clk2, and Clk4, respectively. No inhibitory effect on Clk3, SRPK1, SRPK2, or PKC.

TG003 Chemical Structure

CAS No. 300801-52-9

Selleck's TG003 has been cited by 5 Publications

Purity & Quality Control

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Biological Activity

Description TG003 is a potent and ATP-competitive Cdc2-like kinase (Clk) inhibitor with IC50 of 20 nM, 200 nM, and 15 nM for Clk1, Clk2, and Clk4, respectively. No inhibitory effect on Clk3, SRPK1, SRPK2, or PKC.
Targets
mCLK4 [1]
(Cell-free assay)
mCLK1 [1]
(Cell-free assay)
mCLK2 [1]
(Cell-free assay)
15 nM 20 nM 200 nM
In vitro

TG003 inhibits SF2/ASF-dependent splicing of human β-globin in vitro by suppression of Clk1/Sty-mediated phosphorylation. TG003 inhibits Clk1/Sty kinase activity in mammalian cells, while has no toxic effect on growth of HeLa and COS-7 cells at 10 μM concentration. [1] TG003 blocks IL-1β RNA production by platelets by inhibiting splicing of IL-1β heteronuclear RNA. [2] During 3T3-L1 adipocyte differentiation, TG003 also blocks alternative splicing of PKCβII and expression of PPARγ1 and PPARγ2. [3]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
human STO cells MWTGeY5kfGmxbjDhd5NigQ>? NEX0ZVJOd2S3bHH0bY9vKG:oIFPsb{9{fHlicILlMY1TVkFic4DsbYNqdmdiaX6gbJVu[W5iU2TPJINmdGy|IHHzd4V{e2WmIHHzJINwdmOnboTyZZRqd25icnXxeYlz\WRidH:gbY5kemWjc3WgbY4hdWG2dYLlJGNtczJibWLORUBt\X[nbDDifUBTXC2SQ2KgZY5idHm|aYOsJGVEPTB;Nj62JO69VQ>? NV3ONpRHOjV{MkG2OFk>
Assay
Methods Test Index PMID
Western blot CLK1 / SRPK1 27397683
Immunofluorescence tubulin / emerin 27015110
In vivo TG003 (10 μM) rescues the embryonic defects induced by excessive Clk activity in Xenopus. [1]

Protocol (from reference)

Kinase Assay:

[1]

  • In Vitro Kinase Assay :

    Kinase activity of Clks and SRPKs is assayed in a reaction mixture, containing 200 mM Tris-HCl (pH 7.5), 12.5 mM MgCl2, 8 mM dithiothreitol, 4 mM EGTA, 1–20 μM ATP, 1 μCi of [γ-32P]ATP, 1 μg of synthetic peptide of SF2/ASF RS domain (NH2-RSPSYGRSRSRSRSRSRSRSRSNSRSRSY-OH), and 0.1–1 μg of purified kinases in a final volume of 40 μL. cAMP-dependent protein kinase activity is assayed in a reaction mixture containing 80 mM Tris-HCl (pH 7.5), 12.5 mM MgCl2, 8 mM dithiothreitol, 4 mM EGTA, 10 μM ATP, 1 μCi of [γ-32P]ATP, 5 μg of histone H1, and 1 μg of catalytic subunit of rat cAMP-dependent protein kinase purified. Protein kinase C activity is assayed in a reaction mixture containing 200 mM Tris-HCl (pH 7.5), 12.5 mM MgCl2, 1 mM CaCl2, 80 μg/mL phosphatidylserine, 8 μg/mL diolein, 10 μM ATP, 1 μCi of [γ-32P]ATP, 5 μg of histone H1, and 2 μL of partially purified rat protein kinase C. The final concentration of Me2SO is adjusted to 1% regardless of inhibitor concentration. The reaction mixture is incubated at 30 or 25 °C for mammalian or Xenopus recombinant proteins, respectively, for 10 min, and a half-portion is spotted on P81 phosphocellulose membrane. The kinase assay conditions, including the incubation period and concentration of kinases and substrates, are optimized to maintain the linearity during incubation. The membrane is washed with 5% phosphoric acid solution (SF2/ASF RS domain) or 5% trichloroacetic solution (histone H1) at least over 15 min. The radioactivity is measured using a liquid scintillation counter. The net radioactivity is deduced by subtracting the background count from the reaction mixture without kinase, and the data are expressed as the percentage to the control sample containing the solvent.

Cell Research:

[1]

  • Cell lines: HeLa cells and COS-7 cells
  • Concentrations: ~10 μM
  • Incubation Time: 3 days
  • Method:

    2 × 105 HeLa cells or 1.5 × 105 COS-7 cells resuspended in 2 mL of medium are plated on 6-well dishes, and 2 μL of 10 mM TG003 dissolved in Me2SO (final concentration at 10 μM), or 2 μL of Me2SO, is added to some wells. Cells are trypsinized, and the density is counted every 24 h for 3 days. Cells are then fixed with 1 mL of ice-cold 70% ethanol, washed with PBS, incubated in 1 ml of PBS containing 1 μg/mL DNase-free RNase A and 50 μg/mL propidium iodide for 20 min at 37 °C, and proceeded to cell cycle analysis by FACSCalibur.

Animal Research:

[1]

  • Animal Models: Xenopus laevis embryos
  • Dosages: ~10 μM
  • Administration: --

Solubility (25°C)

In vitro

Chemical Information

Molecular Weight 249.33
Formula

C13H15NO2S

CAS No. 300801-52-9
Storage 3 years -20°C powder
2 years -80°C in solvent
Smiles CCN1C2=C(C=CC(=C2)OC)SC1=CC(=O)C

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Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

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Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

Handling Instructions

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