| Description |
FGFR4-IN-1 is a potent inhibiotr of FGFR4 with IC50 of 0.7 nM.
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| Related Catalog |
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| Target |
FGFR4:0.7 nM (IC50)
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| In Vitro |
FGFR4-IN-1 significantly inhibits the proliferation of HuH-7 hepatocellular carcinoma cells with IC50 of 7.8 nM[1].
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| Kinase Assay |
250000 BaF3-Tel-FGFR4 cells per well are seeded in 96-well tissue culture plates in 40 μL of growth medium supplemented with 10% foetal calf serum, 10 mM HEPES, 1 mM Sodium Pyruvate, 2 mM Stable Glutamine and 1x Penicillin-Streptomycin. Using a liquid handling device, serial 3-fold dilutions of compounds are prepared in DMSO, prediluted in growth medium, followed by transfer of 10 μL/well to the cell plates. After incubation for 1 hour at 37°C/5% CO2, 50 μL of lysis buffer (150 mM NaCl, 20 mM Tris (pH 7.5), 1 mM EDTA, 1 mM EGTA, 1 % Triton X-100, 10 mM NaF, complements with protease inhibitors and phosphatase) inhibitors is added and incubated for 30 minutes on ice with shaking at 300 rpm. Sample plates are then frozen and stored at 70°C. Following thawing on ice, the sample plates are centrifuged for 15 minutes at 1200 rpm at 6°C.
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| Cell Assay |
Methylene blue staining proliferation assay (MBS): The effect of compounds on cell proliferation is assessed using HuH-7 hepatocellular carcinoma cells are cultured in the vendor-recommended medium, 10% foetal calf serum, 1 mM sodium pyruvate, 1x Penicillin/Streptomycin at 37°C in a humidified 5% CO2 incubator. Specifically, 5000 cells/well are seeded in 96-well tissue culture plates in a total media volume of 100 μL/well and increasing compound dilutions or DMSO are added 24 hours thereafter in triplicates. 72 hours after compound addition, cells are fixed by adding 25 and incubated for 10 minutes at room temperature. Cells are washed three times with H2O. Cells are washed 3 times with H2O, 200 mL/well and then lysed by adding 200 mL/well of 3% HCl for 30 minutes at room temperature with shaking. Optical density is measured at A650 nm. The concentration of compound providing 50% of proliferation inhibition with respect to DMSO-treated cells is determined (IC50) using XLFit software.
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| References |
[1]. Ring-fused bicyclic pyridyl derivatives as fgfr4 inhibitors. WO 2015059668 A1
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