5-Methylisoxazole-4-carboxylic acid (4-trifluoromethyl)anilide Arava (TN) 5-methyl-N-[4-(trifluoromethyl)phenyl]-1,2-oxazole-4-carboxamide BDBM50054601 5-Methyl-N-(4-(trifluoromethyl)phenyl)-4-isoxazolecarboxamide CHEMBL960 LEFLUNOMIDE alpha,alpha,alpha-Trifluoro-5-methyl-4-isoxazolecarboxy-p-toluidide
- Giorgis, M; Lolli, ML; Rolando, B; Rao, A; Tosco, P; Chaurasia, S; Marabello, D; Fruttero, R; Gasco, A 1,2,5-Oxadiazole analogues of leflunomide and related compounds. Eur J Med Chem 46: 383-92 (2010)
- Papageorgiou, C; Albert, R; Floersheim, P; Lemaire, M; Bitch, F; Weber, HP; Andersen, E; Hungerford, V; Schreier, MH Pyrazole bioisosteres of leflunomide as B-cell immunosuppressants for xenotransplantation and chronic rejection: scope and limitations. J Med Chem 41: 3530-8 (1998)
- Papagergiou, C; Zurini, M; Weber, HP; Borer, X Leflunomide's Bioactive Metabolite Has the Minimal Structural Requirements for the Efficient Inhibition of Human Dihydroorotate Dehydrogenase Bioorg Chem 25: 233-238 (1997)
- Enzymatic Assay 2 The enzymatic assay couples DHODH activity with bleaching of the dye 2,6-Dichlorophenolindophenol (DCIP) (Knecht and Loffler, 1998; Miller et al., 1968). The assay was conducted in aqueous buffer containing 50 mM Tris, 0.1% Triton X-100, 150 mM potassium chloride, 0.4 μg/mL DHODH, 1 mM dihydroorotate, 0.1 mM decylubiquinone, 0.06 mM DCIP, and 0.17% DMSO at pH 8.0 at room temperature. Compounds were added via pin transfer or via D300 digital dispenser, and the reaction was initiated by addition of substrates. Enzyme activity was monitored kinetically by the reduction in DCIP absorbance at 600 nm. Purified recombinant human DHODH (full-length, C-terminal MYC/DDK-tag) enzyme was purchased from Origene (cat. no. TP039034). Other chemicals, including leflunomide and teriflunomide, were purchased from Sigma-Aldrich. Absorbance measurements were obtained using a Molecular Devices Spectramax M5 plate-reading spectrophotometer.
- Enzymatic Assay Assay 2: The enzymatic assay couples DHODH activity with bleaching of the dye 2,6-Dichlorophenolindophenol (DCIP) (Knecht and Loffler, 1998; Miller et al., 1968). The assay was conducted in aqueous buffer containing 50 mM Tris, 0.1% Triton X-100, 150 mM potassium chloride, 0.4 μg/mL DHODH, 1 mM dihydroorotate, 0.1 mM decylubiquinone, 0.06 mM DCIP, and 0.17% DMSO at pH 8.0 at room temperature. Compounds were added via pin transfer or via D300 digital dispenser, and the reaction was initiated by addition of substrates. Enzyme activity was monitored kinetically by the reduction in DCIP absorbance at 600 nm. Purified recombinant human DHODH (full-length, C-terminal MYC/DDK-tag) enzyme was purchased from Origene (cat. no. TP039034). Other chemicals, including leflunomide and teriflunomide, were purchased from Sigma-Aldrich. Absorbance measurements were obtained using a Molecular Devices Spectramax M5 plate-reading spectrophotometer.
- In Vitro Assay 2 The enzymatic assay couples DHODH activity with bleaching of the dye 2,6-Dichlorophenolindophenol (DCIP) (Knecht and Loffler, 1998; Miller et al. 1968). The assay was conducted in aqueous buffer containing 50 mM Tris, 0.1% Triton X-100, 150 mM potassium chloride, 0.4 μg/mL DHODH, 1 mM dihydroorotate, 0.1 mM decylubiquinone, 0.06 mM DCIP, and 0.17% DMSO at pH 8.0 at room temperature. Compounds were added via pin transfer or via D300 digital dispenser, and the reaction was initiated by addition of substrates. Enzyme activity was monitored kinetically by the reduction in DCIP absorbance at 600 nm. Purified recombinant human DHODH (full-length, C-terminal MYC/DDK-tag) enzyme was purchased from Origene (cat. no. TP039034). Other chemicals, including leflunomide and teriflunomide, were purchased from Sigma-Aldrich. Absorbance measurements were obtained using a Molecular Devices Spectramax M5 plate-reading spectrophotometer.