lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Cc1c(C(=O)N2[C@H]3CC[C@@H]2c2nnc(-c4cscn4)n2C3)cccc1C(F)(F)F
InChIKey: FUEAMKMSQWBPNG-XHDPSFHLSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Ca2+ Flux Assay: 1321N1 cells expressing the recombinant human or rat P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 μl volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250× the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 μL of the compound into 300 μL of assay buffer. A further 3× dilution occurred when transferring 50 μL/well of the compound plate to 100 μL/well in the cell plate. Cells were incubated with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 μL/well of BzATP (final concentration is 250 μM BzATP (human and rat)).
Measurement: Ki = 7.9 nM (pChEMBL 8.100000381469727)
Descriptors: MW 419.43, ALogP 4.09, HBA 6, HBD 0, PSA 63.91, QED 0.63
Compound (SMILES): O=C(c1ccc(Cl)c(F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cscn3)n1C2
InChIKey: LEXDKUWFGJVJTC-QPUJVOFHSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 FLIPR Assay: 1321N1 cells expressing the recombinant human or rat P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 μl volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250× the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 μL of the compound into 300 μL of assay buffer. A further 3× dilution occurred when transferring 50 μL/well of the compound plate to 100 μL/well in the cell plate. Cells were incubated with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 μL/well of BzATP (final concentration is 250 μM BzATP (human and rat)). The fluorescence change was measured 180 seconds after adding the agonist. Peak fluorescence was plotted as a function of BzATP concentration using Origin 7 software.
Measurement: IC50 = 8912 nM (pChEMBL 5.050000190734863)
Descriptors: MW 424.29, ALogP 4.21, HBA 6, HBD 0, PSA 63.91, QED 0.58
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccc(Cl)s3)n1C2
InChIKey: PJMVTHMVIKAXFS-JOYOIKCWSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Ca2+ Flux Assay: 1321N1 cells expressing the recombinant human or rat P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 μl volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250× the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 μL of the compound into 300 μL of assay buffer. A further 3× dilution occurred when transferring 50 μL/well of the compound plate to 100 μL/well in the cell plate. Cells were incubated with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 μL/well of BzATP (final concentration is 250 μM BzATP (human and rat)).
Measurement: Ki = 16 nM (pChEMBL 7.800000190734863)
Descriptors: MW 473.31, ALogP 5.69, HBA 5, HBD 0, PSA 51.02, QED 0.48
Compound (SMILES): CCC(=N\O)/C(C)=C/c1ccc(F)cc1
InChIKey: HKROEBDHHKMNBZ-CHBKHGQFSA-N
Target: Transient receptor potential cation channel subfamily V member 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9HBA0
Assay: Radiometric Calcium Assay: The inhibition of TRPA1 receptor activation was measured as inhibition of allylisothiocyanate (AITC) induced cellular uptake of radioactive calcium. Test compounds were dissolved in DMSO to prepare 10 mM stock solution and then diluted using plain medium with 0.1% BSA and 1.8 mM CaCl2 to get desired concentration. Final concentration of DMSO in the reaction was 0.5% (v/v). Human TRPA1 expressing CHO cells were grown in F-12 DMEM medium with 10% FBS, 1% penicillin-streptomycin solution, and 400 ug/ml of G-418. Cells were seeded 24 h prior to the assay in 96 well plates so as to get 50,000 cells per well on the day of experiment. Cells were treated with test compounds for 10 min followed by addition of AITC at a final concentration of 30 micromolar and 5 uCi/ml 45Ca+2 for 3 min at 25° C. Cells were washed and lysed using buffer containing 1% Triton X-100, 0.1% deoxycholate and 0.1% SDS. Radioactivity in the lysate was measured in Packard Top count.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 207.25, ALogP 3.47, HBA 2, HBD 1, PSA 32.59, QED 0.46
Compound (SMILES): CC12CC3CC(C)(C1)CC(N1[C@@H]4CC[C@H]1C[C@@H](n1c(=O)c(C(=O)O)nc5ccccc51)C4)(C3)C2
InChIKey: DUDXAOUGIMVJPT-JZPJOJOISA-N
Target: Kappa-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41145
Assay: Radioligand Binding Assay: Radioligand dose displacement assays used 0.4-0.8nM [3H]-U69,593 (NEN; 40 Ci/mmole) with 10-20 ug membrane protein (recombinant kappa opioid receptor expressed in HEK 293 cells; in-house prep) in a final volume of 200 uL binding buffer (5% DMSO, 50mM Trizma base, pH 7.4). Non-specific binding was determined in the presence of 10 uM unlabeled naloxone or U69,593. All reactions were performed in 96-well polypropylene plates for 1 h at a temperature of about 25C. Binding reactions were determined by rapid filtration onto 96-well Unifilter GF/C filter plates (Packard) presoaked in 0.5% polyethylenimine (Sigma). Harvesting was performed using a 96-well tissue harvester (Packard) followed by five filtration washes with 200uL ice-cold binding buffer. Filter plates were subsequently dried at 50C for 1-2 hours. Fifty uL/well scintillation cocktail (MicroScint20, Packard) was added and plates were counted in a Packard Top-Count for 1 min/well.
Measurement: Ki = 6866 nM (pChEMBL 5.159999847412109)
Descriptors: MW 461.61, ALogP 5.01, HBA 5, HBD 1, PSA 75.43, QED 0.7
Compound (SMILES): O=C(O)c1nc2ccccc2n([C@H]2C[C@H]3CCC[C@@H](C2)N3C23CC4CC(CC(C4)C2)C3)c1=O
InChIKey: AVCYHJJDUBPVJN-ORRRZOFSSA-N
Target: Delta-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41143
Assay: Radioligand Binding Assay: Radioligand dose-displacement assays used 0.2nM [3H]-Naltrindole (NEN; 33.0 Ci/mmole) with 10-20ug membrane protein (recombinant delta opioid receptor expressend in CHO-K1 cells; Perkin Elmer) in a final volume of 500uL binding buffer (5mM MgCl2, 5% DMSO, 50mM Trizma base, pH 7.4). Non-specific binding was determined in the presence of 25 uM unlabeled naloxone. All reactions were performed in 96-deep well polypropylene plates for 1 h at a temperature of about 25C. Binding reactions were determined by rapid filtration onto 96-well Unifilter GF/C filter plates (Packard) presoaked in 0.5% polyethylenimine (Sigma). Harvesting was performed using a 96-well tissue harvester (Packard) followed by five filtration washes with 500uL ice-cold binding buffer. Filter plates were subsequently dried at 50C for 1-2 hours. Fifty uL/well scintillation cocktail (MicroScint20, Packard) was added and plates were counted in a Packard Top-Count for 1 min/well.
Measurement: Ki = 39396 nM (pChEMBL 4.409999847412109)
Descriptors: MW 447.58, ALogP 4.62, HBA 5, HBD 1, PSA 75.43, QED 0.74
Compound (SMILES): CCNC(=O)C#Cc1ccc2c(c1)NC(=O)/C2=C(\Nc1ccc(CN(C)C)cc1)c1ccccc1
InChIKey: FLBNLJLONKAPLR-DQSJHHFOSA-N
Target: Acyl-CoA dehydrogenase family member 11 (SINGLE PROTEIN), Homo sapiens
UniProt: Q709F0
Assay: Kinobeads (epsilon), multiple immobilized ATP-competitive broad spectrum kinase inhibitors, used to assess residual binding of ~300 proteins simultaneously from cell lysate in the presence of a compound. Quantitative readout performed by mass spectrometry.
Measurement: Kd > 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 464.57, ALogP 4.17, HBA 4, HBD 3, PSA 73.47, QED 0.38
Compound (SMILES): COc1cc(C2=C(c3cn(CCCO)c4ccccc34)C(=O)NC2=O)cc(OC)c1OC
InChIKey: IFUKBECSYNPLCY-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Kinase Assay: The effect of test compounds on the activity of the protein kinases GSK-3β, VEGFR-2 and FLT-3 was evaluated based on half maximal inhibitory concentration (IC50) values determined by Millipore UK Ltd; Gemini Crescent; Dundee Technology Park; Dundee DD2 1SW; UK (IC50Profiler). Detailed protocols can be found at: www.millipore.com/drugdiscovery/dd3/assayprotocols.
Measurement: IC50 = 920 nM (pChEMBL 6.039999961853027)
Descriptors: MW 436.46, ALogP 2.62, HBA 7, HBD 2, PSA 99.02, QED 0.53
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@@H]2CCC[C@H]1c1nnc(-c3ccc(F)cn3)n1C2
InChIKey: KAXAHUBBXCYPAA-WBMJQRKESA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 12.6 nM (pChEMBL 7.900000095367432)
Descriptors: MW 465.84, ALogP 4.9, HBA 5, HBD 0, PSA 63.91, QED 0.5
Compound (SMILES): Cc1cccc(-c2nnc3n2C[C@@H]2CCC[C@H]3N2C(=O)c2cccc(C(F)(F)F)c2Cl)n1
InChIKey: NBZULESLUAZFRE-SUMWQHHRSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 5 nM (pChEMBL 8.300000190734863)
Descriptors: MW 461.88, ALogP 5.07, HBA 5, HBD 0, PSA 63.91, QED 0.53
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CCC[C@@H]1c1nnc(-c3ccccn3)n1C2
InChIKey: GJFWSWDCNPBEIO-BLLLJJGKSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Ca2+ Flux Assay: 1321N1 cells expressing the recombinant human or rat P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 μl volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250× the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 μL of the compound into 300 μL of assay buffer. A further 3× dilution occurred when transferring 50 μL/well of the compound plate to 100 μL/well in the cell plate. Cells were incubated with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 μL/well of BzATP (final concentration is 250 μM BzATP (human and rat)).
Measurement: Ki = 3.2 nM (pChEMBL 8.489999771118164)
Descriptors: MW 447.85, ALogP 4.76, HBA 5, HBD 0, PSA 63.91, QED 0.57
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccc(F)cc3)n1C2
InChIKey: JLCZWQQYVOECLS-XJKSGUPXSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 Radioligand Binding Assay: human or rat P2X7-1321N1 cells were collected and frozen @ −80° C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 μl:10 μl compound (10×)+(b) 40 μl tracer (2.5×)+50 μl membrane (2×). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2008, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4° C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM). The IC50 generated in the binding assay was corrected for tracer concentration and affinity of the tracer to derive at the affinity (Ki) of the test compounds. The data are presented in Tables 1 and 2 under the headings: P2X7 human Ki (μM) and P2X7 rat Ki (μM). Data are analyzed and graphed on Graphpad Prism 5.
Measurement: Ki = 35.5 nM (pChEMBL 7.449999809265137)
Descriptors: MW 450.82, ALogP 5.12, HBA 4, HBD 0, PSA 51.02, QED 0.51
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccccn3)n1C2
InChIKey: YCAGFSLZIXNIAQ-XHDPSFHLSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 1584.9 nM (pChEMBL 5.800000190734863)
Descriptors: MW 433.82, ALogP 4.37, HBA 5, HBD 0, PSA 63.91, QED 0.6
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cnccn3)n1C2
InChIKey: GXIFGAOSXYVTTL-IINYFYTJSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 12.6 nM (pChEMBL 7.900000095367432)
Descriptors: MW 434.81, ALogP 3.77, HBA 6, HBD 0, PSA 76.8, QED 0.61
Compound (SMILES): O=C(c1ccc(F)c(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccccn3)n1C2
InChIKey: XOBAFBCJBBPMMO-IINYFYTJSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 3981.1 nM (pChEMBL 5.400000095367432)
Descriptors: MW 451.81, ALogP 4.51, HBA 5, HBD 0, PSA 63.91, QED 0.54
Compound (SMILES): Cc1c(C(=O)N2[C@H]3CC[C@@H]2c2nnc(-c4ccccn4)n2C3)cccc1C(F)(F)F
InChIKey: BAFAGPLAUVJQTL-SUMWQHHRSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 8491.8 nM (pChEMBL 5.070000171661377)
Descriptors: MW 413.4, ALogP 4.03, HBA 5, HBD 0, PSA 63.91, QED 0.64
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccc(F)cn3)n1C2
InChIKey: PIWFESADTUHGDM-XHDPSFHLSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 12.6 nM (pChEMBL 7.900000095367432)
Descriptors: MW 451.81, ALogP 4.51, HBA 5, HBD 0, PSA 63.91, QED 0.54
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ncccn3)n1C2
InChIKey: UHVGGRPPRZVREM-GXFFZTMASA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 Radioligand Binding Assay: human or rat P2X7-1321N1 cells were collected and frozen @ −80° C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 μl:10 μl compound (10×)+(b) 40 μl tracer (2.5×)+50 μl membrane (2×). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2008, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4° C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM). The IC50 generated in the binding assay was corrected for tracer concentration and affinity of the tracer to derive at the affinity (Ki) of the test compounds. The data are presented in Tables 1 and 2 under the headings: P2X7 human Ki (μM) and P2X7 rat Ki (μM). Data are analyzed and graphed on Graphpad Prism 5.
Measurement: Ki = 6.3 nM (pChEMBL 8.199999809265137)
Descriptors: MW 434.81, ALogP 3.77, HBA 6, HBD 0, PSA 76.8, QED 0.61
Compound (SMILES): O=C(c1ccc(F)c(Cl)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cnccn3)n1C2
InChIKey: MCTLJXIJKPVBNL-TVQRCGJNSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 1923.1 nM (pChEMBL 5.71999979019165)
Descriptors: MW 419.25, ALogP 3.54, HBA 6, HBD 0, PSA 76.8, QED 0.59
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cccnc3)n1C2
InChIKey: VKDZQRGLSAJSNR-SWLSCSKDSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 15.8 nM (pChEMBL 7.800000190734863)
Descriptors: MW 433.82, ALogP 4.37, HBA 5, HBD 0, PSA 63.91, QED 0.6
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cscn3)n1C2
InChIKey: MUGFFAOGIBZAEJ-TVQRCGJNSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 5 nM (pChEMBL 8.300000190734863)
Descriptors: MW 439.85, ALogP 4.43, HBA 6, HBD 0, PSA 63.91, QED 0.59
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ncc(F)cn3)n1C2
InChIKey: PGNGAKNDIHFQII-GXFFZTMASA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 FLIPR Assay: 1321N1 cells expressing the recombinant human or rat P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 μl volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250× the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 μL of the compound into 300 μL of assay buffer. A further 3× dilution occurred when transferring 50 μL/well of the compound plate to 100 μL/well in the cell plate. Cells were incubated with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 μL/well of BzATP (final concentration is 250 μM BzATP (human and rat)). The fluorescence change was measured 180 seconds after adding the agonist. Peak fluorescence was plotted as a function of BzATP concentration using Origin 7 software.
Measurement: IC50 = 6776 nM (pChEMBL 5.170000076293945)
Descriptors: MW 452.8, ALogP 3.91, HBA 6, HBD 0, PSA 76.8, QED 0.55
Compound (SMILES): Cc1cc(-c2nnc3n2C[C@@H]2CC[C@H]3N2C(=O)c2cccc(C(F)(F)F)c2Cl)[nH]n1
InChIKey: JDZGMBZXGAFIMR-IINYFYTJSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 Radioligand Binding Assay: human or rat P2X7-1321N1 cells were collected and frozen @ −80° C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 μl:10 μl compound (10×)+(b) 40 μl tracer (2.5×)+50 μl membrane (2×). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2008, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4° C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM). The IC50 generated in the binding assay was corrected for tracer concentration and affinity of the tracer to derive at the affinity (Ki) of the test compounds. The data are presented in Tables 1 and 2 under the headings: P2X7 human Ki (μM) and P2X7 rat Ki (μM). Data are analyzed and graphed on Graphpad Prism 5.
Measurement: Ki = 7.9 nM (pChEMBL 8.100000381469727)
Descriptors: MW 436.83, ALogP 4.01, HBA 5, HBD 1, PSA 79.7, QED 0.66
Compound (SMILES): O=C(c1ccc(F)c(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cscn3)n1C2
InChIKey: KQVQQNCOTWFYGO-QPUJVOFHSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 Radioligand Binding Assay: human or rat P2X7-1321N1 cells were collected and frozen @ −80° C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 μl:10 μl compound (10×)+(b) 40 μl tracer (2.5×)+50 μl membrane (2×). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2008, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4° C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM). The IC50 generated in the binding assay was corrected for tracer concentration and affinity of the tracer to derive at the affinity (Ki) of the test compounds. The data are presented in Tables 1 and 2 under the headings: P2X7 human Ki (μM) and P2X7 rat Ki (μM). Data are analyzed and graphed on Graphpad Prism 5.
Measurement: Ki = 6.3 nM (pChEMBL 8.199999809265137)
Descriptors: MW 457.84, ALogP 4.57, HBA 6, HBD 0, PSA 63.91, QED 0.53
Compound (SMILES): Cc1c(C(=O)N2[C@H]3CC[C@@H]2c2nnc(-c4cscn4)n2C3)cccc1C(F)(F)F
InChIKey: FUEAMKMSQWBPNG-XHDPSFHLSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 Radioligand Binding Assay: human or rat P2X7-1321N1 cells were collected and frozen @ −80° C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 μl:10 μl compound (10×)+(b) 40 μl tracer (2.5×)+50 μl membrane (2×). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2008, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4° C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM). The IC50 generated in the binding assay was corrected for tracer concentration and affinity of the tracer to derive at the affinity (Ki) of the test compounds. The data are presented in Tables 1 and 2 under the headings: P2X7 human Ki (μM) and P2X7 rat Ki (μM). Data are analyzed and graphed on Graphpad Prism 5.
Measurement: Ki = 7.9 nM (pChEMBL 8.100000381469727)
Descriptors: MW 419.43, ALogP 4.09, HBA 6, HBD 0, PSA 63.91, QED 0.63
Compound (SMILES): O=C(c1ccc(Cl)c(F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cscn3)n1C2
InChIKey: LEXDKUWFGJVJTC-QPUJVOFHSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 8912.5 nM (pChEMBL 5.050000190734863)
Descriptors: MW 424.29, ALogP 4.21, HBA 6, HBD 0, PSA 63.91, QED 0.58
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccc(Cl)s3)n1C2
InChIKey: PJMVTHMVIKAXFS-JOYOIKCWSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 Radioligand Binding Assay: human or rat P2X7-1321N1 cells were collected and frozen @ −80° C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 μl:10 μl compound (10×)+(b) 40 μl tracer (2.5×)+50 μl membrane (2×). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2008, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4° C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM). The IC50 generated in the binding assay was corrected for tracer concentration and affinity of the tracer to derive at the affinity (Ki) of the test compounds. The data are presented in Tables 1 and 2 under the headings: P2X7 human Ki (μM) and P2X7 rat Ki (μM). Data are analyzed and graphed on Graphpad Prism 5.
Measurement: Ki = 15.8 nM (pChEMBL 7.800000190734863)
Descriptors: MW 473.31, ALogP 5.69, HBA 5, HBD 0, PSA 51.02, QED 0.48
Compound (SMILES): CCC(=N\O)/C(C)=C/c1ccc(F)cc1
InChIKey: HKROEBDHHKMNBZ-CHBKHGQFSA-N
Target: Transient receptor potential cation channel subfamily M member 8 (SINGLE PROTEIN), Homo sapiens
UniProt: Q7Z2W7
Assay: Radiometric Calcium Assay: The inhibition of TRPA1 receptor activation was measured as inhibition of allylisothiocyanate (AITC) induced cellular uptake of radioactive calcium. Test compounds were dissolved in DMSO to prepare 10 mM stock solution and then diluted using plain medium with 0.1% BSA and 1.8 mM CaCl2 to get desired concentration. Final concentration of DMSO in the reaction was 0.5% (v/v). Human TRPA1 expressing CHO cells were grown in F-12 DMEM medium with 10% FBS, 1% penicillin-streptomycin solution, and 400 ug/ml of G-418. Cells were seeded 24 h prior to the assay in 96 well plates so as to get 50,000 cells per well on the day of experiment. Cells were treated with test compounds for 10 min followed by addition of AITC at a final concentration of 30 micromolar and 5 uCi/ml 45Ca+2 for 3 min at 25° C. Cells were washed and lysed using buffer containing 1% Triton X-100, 0.1% deoxycholate and 0.1% SDS. Radioactivity in the lysate was measured in Packard Top count.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 207.25, ALogP 3.47, HBA 2, HBD 1, PSA 32.59, QED 0.46
Compound (SMILES): CC12CC3CC(C)(C1)CC(N1[C@@H]4CC[C@H]1C[C@@H](n1c(=O)c(C(=O)O)nc5ccccc51)C4)(C3)C2
InChIKey: DUDXAOUGIMVJPT-JZPJOJOISA-N
Target: Delta-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41143
Assay: Radioligand Binding Assay: Radioligand dose-displacement assays used 0.2nM [3H]-Naltrindole (NEN; 33.0 Ci/mmole) with 10-20ug membrane protein (recombinant delta opioid receptor expressend in CHO-K1 cells; Perkin Elmer) in a final volume of 500uL binding buffer (5mM MgCl2, 5% DMSO, 50mM Trizma base, pH 7.4). Non-specific binding was determined in the presence of 25 uM unlabeled naloxone. All reactions were performed in 96-deep well polypropylene plates for 1 h at a temperature of about 25C. Binding reactions were determined by rapid filtration onto 96-well Unifilter GF/C filter plates (Packard) presoaked in 0.5% polyethylenimine (Sigma). Harvesting was performed using a 96-well tissue harvester (Packard) followed by five filtration washes with 500uL ice-cold binding buffer. Filter plates were subsequently dried at 50C for 1-2 hours. Fifty uL/well scintillation cocktail (MicroScint20, Packard) was added and plates were counted in a Packard Top-Count for 1 min/well.
Measurement: Ki = 16831 nM (pChEMBL 4.769999980926514)
Descriptors: MW 461.61, ALogP 5.01, HBA 5, HBD 1, PSA 75.43, QED 0.7
Compound (SMILES): CCNC(=O)C#Cc1ccc2c(c1)NC(=O)/C2=C(\Nc1ccc(CN(C)C)cc1)c1ccccc1
InChIKey: FLBNLJLONKAPLR-DQSJHHFOSA-N
Target: Very long-chain specific acyl-CoA dehydrogenase, mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: P49748
Assay: Kinobeads (epsilon), multiple immobilized ATP-competitive broad spectrum kinase inhibitors, used to assess residual binding of ~300 proteins simultaneously from cell lysate in the presence of a compound. Quantitative readout performed by mass spectrometry.
Measurement: Kd > 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 464.57, ALogP 4.17, HBA 4, HBD 3, PSA 73.47, QED 0.38
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@@H]2CCC[C@H]1c1nnc(-c3ccc(F)cn3)n1C2
InChIKey: KAXAHUBBXCYPAA-WBMJQRKESA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 13.3 nM (pChEMBL 7.880000114440918)
Descriptors: MW 465.84, ALogP 4.9, HBA 5, HBD 0, PSA 63.91, QED 0.5
Compound (SMILES): Cc1cccc(-c2nnc3n2C[C@@H]2CCC[C@H]3N2C(=O)c2cccc(C(F)(F)F)c2Cl)n1
InChIKey: NBZULESLUAZFRE-SUMWQHHRSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: FLIPR Assay: Ca.sup.2+flux: 1321N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 .mu.l volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl.sub.2, 1 MgCl.sub.2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250.times. the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 .mu.L of the compound into 300 .mu.L of assay buffer. A further 3.times. dilution occurred when transferring 50 .mu.L/well of the compound plate to 100 .mu.L/well in the cell plate.
Measurement: IC50 = 56 nM (pChEMBL 7.25)
Descriptors: MW 461.88, ALogP 5.07, HBA 5, HBD 0, PSA 63.91, QED 0.53
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CCC[C@@H]1c1nnc(-c3ccccn3)n1C2
InChIKey: GJFWSWDCNPBEIO-BLLLJJGKSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Ca2+ Flux Assay: 1321N1 cells expressing the recombinant human or rat P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 μl volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250× the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 μL of the compound into 300 μL of assay buffer. A further 3× dilution occurred when transferring 50 μL/well of the compound plate to 100 μL/well in the cell plate. Cells were incubated with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 μL/well of BzATP (final concentration is 250 μM BzATP (human and rat)).
Measurement: Ki = 6.3 nM (pChEMBL 8.199999809265137)
Descriptors: MW 447.85, ALogP 4.76, HBA 5, HBD 0, PSA 63.91, QED 0.57
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccc(F)cc3)n1C2
InChIKey: JLCZWQQYVOECLS-XJKSGUPXSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z1M0
Assay: P2X7 FLIPR Assay: 1321N1 cells expressing the recombinant human or rat P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 μl volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250× the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 μL of the compound into 300 μL of assay buffer. A further 3× dilution occurred when transferring 50 μL/well of the compound plate to 100 μL/well in the cell plate. Cells were incubated with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 μL/well of BzATP (final concentration is 250 μM BzATP (human and rat)). The fluorescence change was measured 180 seconds after adding the agonist. Peak fluorescence was plotted as a function of BzATP concentration using Origin 7 software.
Measurement: IC50 = 10000 nM (pChEMBL 5)
Descriptors: MW 450.82, ALogP 5.12, HBA 4, HBD 0, PSA 51.02, QED 0.51
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccccn3)n1C2
InChIKey: YCAGFSLZIXNIAQ-XHDPSFHLSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 12.6 nM (pChEMBL 7.900000095367432)
Descriptors: MW 433.82, ALogP 4.37, HBA 5, HBD 0, PSA 63.91, QED 0.6
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3cnccn3)n1C2
InChIKey: GXIFGAOSXYVTTL-IINYFYTJSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 7.9 nM (pChEMBL 8.100000381469727)
Descriptors: MW 434.81, ALogP 3.77, HBA 6, HBD 0, PSA 76.8, QED 0.61
Compound (SMILES): O=C(c1ccc(F)c(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccccn3)n1C2
InChIKey: XOBAFBCJBBPMMO-IINYFYTJSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 13.8 nM (pChEMBL 7.860000133514404)
Descriptors: MW 451.81, ALogP 4.51, HBA 5, HBD 0, PSA 63.91, QED 0.54
Compound (SMILES): O=C(c1cccc(C(F)(F)F)c1Cl)N1[C@H]2CC[C@@H]1c1nnc(-c3ccc(F)cn3)n1C2
InChIKey: PIWFESADTUHGDM-XHDPSFHLSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
Assay: Radioligand Binding: Human or rat P2X7-1321N1 cells were collected and frozen @-80.degree. C. On the day of the experiment, cell membrane preparations were made according to standard published methods. The total assay volume was 100 .mu.l:10 .mu.l compound (10.times.)+(b) 40 .mu.l tracer (2.5.times.)+50 .mu.l membrane (2.times.). The tracer used for the assay was tritiated A-804598. The compound can be prepared as described in the literature. (Donnelly-Roberts, D. Neuropharmacology 2009, 56 (1), 223-229.) Compounds, tracer and membranes were incubated for 1 hour @ 4.degree. C. The assay was terminated by filtration (GF/B filters pre-soaked with 0.3% PEI) and washed with washing buffer (Tris-HCl 50 mM).
Measurement: Ki = 31.6 nM (pChEMBL 7.5)
Descriptors: MW 451.81, ALogP 4.51, HBA 5, HBD 0, PSA 63.91, QED 0.54