ABBV-318 [Ligand Id: 12002] activity data from GtoPdb and ChEMBL

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ChEMBL ligand: CHEMBL3970885
  • Nav1.7/Sodium channel protein type IX alpha subunit in Human [ChEMBL: CHEMBL4296] [GtoPdb: 584] [UniProtKB: Q15858]
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  • Nav1.8/Sodium channel protein type X alpha subunit in Human [ChEMBL: CHEMBL5451] [GtoPdb: 585] [UniProtKB: Q9Y5Y9]
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DB Assay description Assay Type Standard value Standard parameter Original value Original units Original parameter Reference
Nav1.7/Sodium channel protein type IX alpha subunit in Human (target type: SINGLE PROTEIN) [ChEMBL: CHEMBL4296] [GtoPdb: 584] [UniProtKB: Q15858]
ChEMBL FRET-Based Membrane Potential Assay: Two days prior to the experiment, frozen HEK293 cells stably expressing recombinant human Nav1.7 were quickly thawed and plated at 25,000 cells/well in growth medium [DMEM (Invitrogen #11965) with 10% HI FBS (Invitrogen #10082), 2 mM glutamine, 100 units/mL penicillin, 0.1 mg/mL streptomycin (PSG, Sigma #G1146), and 500 ug/mL Geneticin (Invitrogen #10131)] in black-walled, clear-bottom 384-well poly-D-lysine-coated assay plates (Greiner Bio-One, Frickenhausen, Germany) and incubated in a humidified 5% CO2 incubator at 37° C. On the day of the assay, medium was removed by aspiration, and cells were washed with assay buffer [HBSS (Invitrogen, Carlsbad, Calif.) containing 20 mM HEPES (Invitrogen, Carlsbad, Calif.)]. After washing, 30 uL assay buffer containing the fluorescent voltage-sensor probe CC2-DMPE (Invitrogen, Carlsbad, Calif.) at 20 uM and 0.01% pluronic F-127 (Invitrogen, Carlsbad, Calif.) was added to the cells. Cells were incubated for 40 minutes at room temperature in the dark. Following the incubation, the cells were washed and 30 uL assay buffer containing 2.5 uM DiSBAC2(3) substrate (Invitrogen, Carlsbad, Calif.) and 0.5 mM VABSC-1 (Invitrogen, Carlsbad, Calif.) was added to the cells. The cells were incubated for 90 minutes at room temperature in the dark. Fluorescence readings were made using a FLIPR TETRA (Molecular Devices, Sunnyvale Calif.) equipped with voltage-sensor probe optics. At the start of each experiment the optimal (EC80) concentration of depolarizing agent (veratridine) was determined by testing a dilution curve of assay buffer containing veratridine (Sigma-Aldrich, St. Louis, Mo.) and 1 mg/mL scorpion venom (SVqq, from Leiurus quinquestriatus; Sigma-Aldrich, St. Louis, Mo.). Compounds were dissolved in dimethyl sulfoxide, and 8-point, 1:3 dilution concentration-response curves were prepared in duplicate in dimethyl sulfoxide, followed by preparation of 0.8 uL/well daughter plates of the dilutions. Test compounds in the daughter plate were diluted to (Ë¿3x) solutions in assay buffer immediately before assaying. Using the FLIPR TETRA, 20 uL of the (3x) compound solutions were first added to the cells, then 20 uL of depolarizing solution (3xEC80 veratridine+SVqq) were added 3 minutes later to activate the channel. Changes in fluorescence were measured at wavelengths of 440-480 nm and 565-625 nm over the course of the experimental run. Membrane depolarization was expressed as a ratio of the maximum F440-480 nm/F565-625 nm reading above average baseline F440-480 nm/F565-625 nm reading. IC50 values were calculated from curve fits of the ratio data using a four-parameter logistic Hill equation (Accelrys Assay Explorer 3.3 Client, Accelrys, San Diego, Calif.) with percent inhibition plotted against compound concentration. B 5.53 pIC50 2980 nM IC50 US-9452986-B2. 6-heteroaryloxy- or 6-aryloxy-quinoline-2-carboxamides and method of use (2016)
GtoPdb Determined in a FRET assay - 5.55 pIC50 2800 nM IC50 Bioorg Med Chem (2022) 63: 116743 [PMID:35436748]
Nav1.8/Sodium channel protein type X alpha subunit in Human (target type: SINGLE PROTEIN) [ChEMBL: CHEMBL5451] [GtoPdb: 585] [UniProtKB: Q9Y5Y9]
ChEMBL FRET-Based Membrane Potential Assay: Two days prior to the experiment, frozen HEK293 cells stably expressing recombinant human Nav1.8 (Essen, Ann Arbor, Mich.) were quickly thawed and plated at 22,500 cells/well in growth medium [MEM (Invitrogen #11095) with 10% FBS (Invitrogen #10082), 1 mM sodium pyruvate (Invitrogen, #C11360), 10 units/mL penicillin 10 U/mL streptomycin 29.2 ug/mL glutamine ((PSG 1%, Invitrogen #10378), 400 ug/mL zeocin (Invitrogen #R250) in black-walled, clear-bottom 384-well poly-D-lysine-coated assay plates (Greiner Bio-One, Frickenhausen, Germany) and incubated in a humidified 5% CO2 incubator at 37° C. On the day of the assay, medium was removed by aspiration, and cells were washed with assay buffer [HBSS (Invitrogen, Carlsbad, Calif.) containing 20 mM HEPES (Invitrogen, Carlsbad, Calif.)]. After washing, 30 uL assay buffer containing the fluorescent voltage-sensor probe CC2-DMPE (Invitrogen, Carlsbad, Calif.) at 20 uM and 0.01% pluronic F-127 (Invitrogen, Carlsbad, Calif.) was added to the cells. Cells were incubated for 40 minutes at room temperature in the dark. Following the incubation, the cells were washed and 30 uL assay buffer containing 2.5 uM DiSBAC2(3) substrate (Invitrogen, Carlsbad, Calif.) and 0.5 mM VABSC-1 (Invitrogen, Carlsbad, Calif.) was added to the cells. The cells were incubated for 60 minutes at room temperature in the dark. Fluorescence readings were made using a FLIPR TETRA(Molecular Devices, Sunnyvale Calif.) equipped with voltage-sensor probe optics. The depolarizing agent, veratridine (Sigma-Aldrich, St. Louis, Mo.), was made up at 3ÿ concentrations in assay buffer containing 1 mg/mL scorpion venom (SVqq, from Leiurus quinquestriatus; Sigma-Aldrich, St. Louis, Mo.). B 5.41 pIC50 3850 nM IC50 US-9452986-B2. 6-heteroaryloxy- or 6-aryloxy-quinoline-2-carboxamides and method of use (2016)
GtoPdb Determined in a FRET assay - 5.42 pIC50 3800 nM IC50 Bioorg Med Chem (2022) 63: 116743 [PMID:35436748]

ChEMBL data shown on this page come from version 28:

Gaulton A, Hersey A, Nowotka M, Bento AP, Chambers J, Mendez D, Mutowo P, Atkinson F, Bellis LJ, Cibrián-Uhalte E, Davies M, Dedman N, Karlsson A, Magariños MP, Overington JP, Papadatos G, Smit I, Leach AR. (2017) 'The ChEMBL database in 2017.' Nucleic Acids Res., 45(D1). DOI: 10.1093/nar/gkw1074. [PMCID:5210557]