lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Cc1ccc(-n2nc(C(C)(C)C)cc2NC(=O)Nc2cnc(Oc3ccc(-c4cnc(N5CCCC5=O)s4)cc3)nc2)cc1C
InChIKey: AJHPWAKXLXZBRY-UHFFFAOYSA-N
Target: High affinity nerve growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P04629
Assay: LanthaScreen Kinase Assay: TrkA-inhibiting activity was measured according to the following method using LanthaScreen kinase assay reagents (Invitrogen; Fluorescein-Poly GT, LanthaScreen Tb-PY20 and TR-FRET Dilution Buffer), TrkA (Invitrogen), ATP (Sigma-Aldrich), a kinase reaction buffer (50 mM HEPES buffer (Sigma-Aldrich) (pH 7.5) containing 0.01% Brij35 (Sigma-Aldrich), 10 mM MgCl2 (Sigma-Aldrich) and 1 mM EGTA (Sigma-Aldrich)) and 0.5 M EDTA (pH 8.0) (Invitrogen).The present compound was dissolved in dimethylsulfoxide (hereinafter abbreviated as DMSO) to prepare a 10 mM solution. The test compound at 10 mM was distributed into a 96-well plate and serially diluted with DMSO with the geometrical ratio of 3. The serial dilutions (DMSO solutions) were diluted with the kinase reaction buffer to 20-fold to prepare solutions of the present compound with 5-times concentrations (DMSO concentration: 5%). TrkA was diluted with the kinase reaction buffer to prepare a solution at 38.5 ng/mL (enzyme solution). Adenosine
Measurement: IC50 = 80 nM (pChEMBL 7.099999904632568)
Descriptors: MW 622.76, ALogP 7.26, HBA 9, HBD 2, PSA 127.16, QED 0.2
Compound (SMILES): CCN(c1cc(Cl)cc(C(=O)NCc2c(C)cc(C)[nH]c2=O)c1C)C1CCC(C(C)C)CC1
InChIKey: NZONDTYRDVXWID-UHFFFAOYSA-N
Target: Histone-lysine N-methyltransferase EZH2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q15910
Assay: Inhibition Assay: Test compounds were serially diluted 3-fold in DMSO in a 10 point-curve and 1 uL was spotted into a 384-well microplate in duplicate using a Platemate Plus equipped with 384-channel head (Thermo Scientific). The final top concentration of test compounds in the assay was 10 uM. Positive control (100% inhibition standard) was 1 mM final concentration of SAH and negative control (0% inhibition standard) contained 1 uL of DMSO. Test compounds were then incubated for 30 minutes with 40 uL per well of wild-type EZH2 (final concentration was 4 nM), Y641F EZH2 (final concentration was 0.1 nM) and A677G and A687V EZH2 (for each, final concentration was 2nM) and peptide in 1x assay buffer (20 mM BICINE pH =7.6, 1 mM DTT, 0.002% TWEEN 20 (generic: polysorbate 20), 0.005% BSG). For the wild-type EZH2 and A677G EZH2 assays, biotinylated peptide H3:21-44 with unmethylated K27 was present at a final concentration of 200 nM, while in the A687V EZH2 assay, biotinylated peptide H3.
Measurement: IC50 = 30 nM (pChEMBL 7.519999980926514)
Descriptors: MW 472.07, ALogP 5.92, HBA 3, HBD 2, PSA 65.2, QED 0.53
Compound (SMILES): CC1CCCN1c1cccc(Nc2cc(-c3ccc4cn[nH]c4c3)nn3ccnc23)n1
InChIKey: WEVYXRIMMPJRKN-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Inhibition Assay: Assay plates: 96-well MultiScreen 0.65 um filter plates (Millipore Cat. No.: MADVNOB10)Streptavidin coated beads: Streptavidin Sepharose, suspension 5.0 mL, in 50 mM EDTA/PBS diluted (1:100), (Amersham, Cat. No.: 17-5113-01)Compounds: 10 mM in 100% dimethylsulfoxide (DMSO), final conc.: compound 0.003-100 uM in 10% DMSOEnzyme: SYK RPA purified, truncated construct of Spleen Tyrosine Kinase aa 360-635, stock solution 1 mg/mL, MW: 31.2 KDa, final conc.:0.0005 uM. Peptide 1: biotinylated peptide is derived from a naturally occurring phosphor-acceptor consensus sequence (Biotin-EPEGDYEEVLE), special order from QCB, stock solution 20 mM, final conc.: 5.0 uM.ATP: Adenosine-5'-triphosphate 20 mM, (ROCHE Cat. No.: 93202720), final concentration: 20 uM Buffer: HEPES: 2-Hydroxyethyl piperazine-2-ethanesulfonic acid (Sigma, Cat. No.: H-3375)final concentration: 50 mM HEPES pH7.5BSA: Bovine Serum Albumin Fraction V, fatty acid free (Roche Diagnostics GmbH, Cat. No. 9100221).
Measurement: IC50 = 75 nM (pChEMBL 7.119999885559082)
Descriptors: MW 410.49, ALogP 4.4, HBA 7, HBD 2, PSA 87.03, QED 0.46
Compound (SMILES): CC(C)CNC(=O)C[C@H]1C[C@@]2(O)[C@H]3Cc4ccc(O)cc4[C@@]2(CCN3C)CC1=O
InChIKey: CUZMLGRZQOZVGB-DQOBCGMHSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: [35S]GTPγS Functional Assay: [35S]GTPγS functional assays were conducted using freshly thawed μ-receptor membranes prepared from a cell line expressing recombinant μ opioid receptor in a HEK-293, CHO or U-2 OS cell background or purchased from a commercial source (Perkin Elmer, Shelton, Conn.; or DiscovRx, Fremont, Calif.). Assay reactions were prepared by sequentially adding the following reagents to binding buffer (100 mM NaCl, 10 mM MgCl2, 20 mM HEPES, pH 7.4) on ice (final concentrations indicated): membrane protein (0.026 mg/mL), saponin (10 mg/mL), GDP (3 mM) and [35S]GTPγS (0.20 nM; Perkin Elmer, Shelton, Conn.). The prepared membrane solution (190 μl/well) was transferred to 96-shallow well polypropylene plates containing 10 μl of 20× concentrated stock solutions of the agonist [D-Ala2, N-methyl-Phe4 Gly-ol5]-enkephalin (DAMGO) prepared in dimethyl sulfoxide (DMSO). Plates were incubated for 30 min at about 25° C. with shaking. Reactions were terminated by rapid filtration onto 96-well Unifilter GF/B filter plates (Perkin Elmer, Shelton, Conn.) using a 96-well tissue harvester (Perkin Elmer, Shelton, Conn.) followed by three filtration washes with 200 μl of ice-cold wash buffer (10 mM NaH2PO4, 10 mM Na2HPO4, pH 7.4). Filter plates were subsequently dried at 50° C. for 2-3 hours. BetaScint scintillation cocktail (Perkin Elmer, Shelton, Conn.) was added (50 μl/well) and plates were counted using a Packard Top-Count for 1 min/well.
Measurement: EC50 = 2331 nM (pChEMBL 5.630000114440918)
Descriptors: MW 400.52, ALogP 1.76, HBA 5, HBD 3, PSA 89.87, QED 0.72
Compound (SMILES): CC(C)(C)CNC(=O)C1CCN(Cc2cccc(NC(=O)c3cccc(C(F)(F)F)n3)c2)CC1
InChIKey: ONWLGJNVCYUZKE-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 6 nM (pChEMBL 8.220000267028809)
Descriptors: MW 476.54, ALogP 4.73, HBA 4, HBD 2, PSA 74.33, QED 0.63
Compound (SMILES): CN1CCN(Cc2ccc(-c3cn4c5nc(ncc35)NCCCCCCCCNC(=O)[C@@H](N)CCCC4)cc2)CC1
InChIKey: DKZQQJXYLNUWFT-LJAQVGFWSA-N
Target: Tyrosine-protein kinase Mer (SINGLE PROTEIN), Homo sapiens
UniProt: Q12866
Assay: Inhibition of human MerTK kinase domain (1585 to 3000 residues) expressed in HEK293 cells co-expressing rat EGFR LBD assessed as inhibition of EGF-stimulated MerTK phosphorylation preincubated for 30 mins followed by EGF stimulation for 15 mins by ELISA
Measurement: EC50 = 160 nM (pChEMBL 6.800000190734863)
Descriptors: MW 560.79, ALogP 4.23, HBA 8, HBD 3, PSA 104.34, QED 0.44
Compound (SMILES): COc1cc(OC)cc(N(CCNS(=O)(=O)C(C)C)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: ZAPUSNWRFJVSGX-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 12.59 nM (pChEMBL 7.900000095367432)
Descriptors: MW 510.62, ALogP 3.51, HBA 9, HBD 1, PSA 111.47, QED 0.35
Compound (SMILES): Cc1ccc(OCC(=O)O)c(C)c1NCc1cc(-c2cccc(F)c2)ccc1F
InChIKey: CLMWSKDETXLTLV-UHFFFAOYSA-N
Target: Prostaglandin E2 receptor EP2 subtype (SINGLE PROTEIN), Homo sapiens
UniProt: P43116
Assay: Cellular Functional Assay: Human CHO cells expressing the recombinant human prostanoid EP2 receptor were suspended in assay medium (HBSS buffer (Invitrogen) containing 20 mM HEPES (pH7.4) and 500 μM isobutyl-methylxanthine IBMX) and plated out to yield approximately 1×104 cells/well in a 96-well. Following this, cells were incubated with agonists for 30 min in the presence of the test compounds. For stimulated control measurements, separate assay wells contain 10 μM PGE2 (control specific agonist). All incubations were carried out at 37° C. in a 5% CO2 atmosphere. Following incubation, the amount of cAMP in each well was determined by HTRF method. The cells are lysed and the fluorescence acceptor (D2-labeled cAMP) and fluorescence donor (anti-cAMP antibody labeled with europium cryptate) are added.
Measurement: EC50 = 26 nM (pChEMBL 7.579999923706055)
Descriptors: MW 397.42, ALogP 5.32, HBA 3, HBD 2, PSA 58.56, QED 0.57
Compound (SMILES): COc1cccc(OC)c1C1CCCC(=O)N1Cc1cnn(-c2ccccc2)c1
InChIKey: LMMPJOIAPQATRY-UHFFFAOYSA-N
Target: Orexin/Hypocretin receptor type 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O43613
Assay: In Vitro Assay: Intracellular Calcium Measurements : In Vitro Assay: Intracellular Calcium Measurements:Chinese hamster ovary (CHO) cells expressing the human orexin-1 receptor and the human orexin-2 receptor, respectively, are grown in culture medium (Ham F-12 with L-Glutamine) containing 300 g/ml G418, 100 U/ml penicillin, 100 g/ml streptomycin and 10% heat inactivated fetal calf serum (FCS). The cells are seeded at 20,000 cells/well into 384-well black clear bottom sterile plates (Greiner). The seeded plates are incubated overnight at 37° C. in 5% CO2. Human orexin-A as an agonist is prepared as 1 mM stock solution in MeOH:water (1:1), diluted in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/l and 20 mM HEPES for use in the assay at a final concentration of 3 nM.Antagonists are prepared as 10 mM stock solution in DMSO, then diluted in 384-well plates using DMSO followed by a transfer of the dilutions into in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/l and 20 mM HEPES. On the day of the assay, 50 μl of staining buffer (HBSS containing 1% FCS, 20 mM HEPES, NaHCO3: 0.375 g/l, 5 mM probenecid (Sigma) and 3 M of the fluorescent calcium indicator fluo-4 AM (1 mM stock solution in DMSO, containing 10% pluronic) is added to each well. The 384-well cell-plates are incubated for 50 min at 37° C. in 5% CO2 followed by equilibration at RT for 30 min before measurement.
Measurement: IC50 = 167 nM (pChEMBL 6.78000020980835)
Descriptors: MW 391.47, ALogP 4.14, HBA 5, HBD 0, PSA 56.59, QED 0.63
Compound (SMILES): CC[C@H](N)CNc1nnc(C(N)=O)c(Nc2cccc3sccc23)n1
InChIKey: CVZHLQQUVFDYRI-VIFPVBQESA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Syk Kinase Inhibition Assay: The in vitro inhibitory activity of the compound of the present invention against Syk kinase activity was assayed under the following conditions: the purified human Syk protein used in the test were purchased from Carna Biosciences, Inc. For the inhibitory activity assay on the compound, the compound of the present invention was first serially diluted with dimethyl sulfoxide (DMSO). Next, the purified human Syk protein FL-Peptide 22 (final concentration: 1 uM) (Caliper Life Sciences, Inc.), magnesium chloride (final concentration: 5 mM), ATP (final concentration: 30 uM), and each DMSO solution of the compound of the present invention (final concentration of DMSO: 5%) were added into a buffer solution for reaction (20 mM tris-HCl pH 7.5, 20 mM NaCl, 2.5 mM DTT, 0.02% Tween-20, 1% glycerol, 0.01 mM Pefabloc), and the mixture was then incubated at 25° C. for 30 minutes to perform kinase reaction. The kinase reaction was stopped by the addition thereto of EDTA.
Measurement: IC50 = 0.35 nM (pChEMBL 9.460000038146973)
Descriptors: MW 357.44, ALogP 2.08, HBA 8, HBD 4, PSA 131.84, QED 0.51
Compound (SMILES): COCc1nc2c(C(=O)Nc3ccc4[nH]ccc4c3)cc(NC(=O)c3ccccc3C(F)(F)F)cc2[nH]1
InChIKey: ZROXIEHEELLDRF-UHFFFAOYSA-N
Target: Prostaglandin E synthase (SINGLE PROTEIN), Homo sapiens
UniProt: O14684
Assay: Inhibition Assay: mPGES-1 microsome fractions were prepared from CHO-K1 cells transiently transfected with plasmid encoding the human mPGES-1cDNA. Microsomes were diluted with potassium phosphate buffer containing reduced glutathione (pH7.4), and DMSO containing test compound or DMSO alone was added (such that DMSO final concentration would be 1% in each) and incubated at 4° C. for 20 minutes. Then, the enzymatic reactions were initiated by the addition of PGH2 substrate (final concentration 1 μM) and incubated at 4° C. for 60 seconds. The reaction was terminated by the addition of a citrate solution (final citrate concentration 50 mM) containing ferric chloride (final concentration 1 mg/mL). PGE2 production in the enzyme reaction aliquot was measured using HTRF kit (Cisbio International, catalogue #62P2APEC).
Measurement: IC50 = 26.5 nM (pChEMBL 7.579999923706055)
Descriptors: MW 507.47, ALogP 5.71, HBA 4, HBD 4, PSA 111.9, QED 0.24
Compound (SMILES): CCC(=O)Nc1cc2c(Nc3ccc(OCc4ccncc4)cc3)ncnc2cc1OC
InChIKey: MFPXZVLJHPMMPW-UHFFFAOYSA-N
Target: Receptor tyrosine-protein kinase erbB-4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q15303
Assay: Irreversible inhibition of GST-tagged ERBB4 (unknown origin) (Gly-259 to Gly-690 residues) expressed in baculovirus infected Sf9 insect cells assessed as reduction in Glu/Tyr copolymer phosphorylation after 6 mins by ELISA
Measurement: IC50 = 428 nM (pChEMBL 6.369999885559082)
Descriptors: MW 429.48, ALogP 4.7, HBA 7, HBD 2, PSA 98.26, QED 0.42
Compound (SMILES): COc1cccc(NC(=O)Nc2ccc(C3CN=C4c5ccccc5NC(=O)N4C3)cc2)c1
InChIKey: SCMILNDAVJFHIK-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Kinase Assay: Biochemical KDR kinase assays were performed in 96 well microtiter plates that were coated overnight with 75 μg/well of poly-Glu-Tyr (4:1) in 10 mM Phosphate Buffered Saline (PBS), pH 7.4. The coated plates were washed with 2 mLs per well PBS+0.05% Tween-20 (PBS-T), blocked by incubation with PBS containing 1% BSA, then washed with 2 mLs per well PBS-T prior to starting the reaction. Reactions were carried out in 100 μL reaction volumes containing 2.7 μM ATP in kinase buffer (50 mM Hepes buffer pH 7.4, 20 mM MgCl2, 0.1 mM MnCl2 and 0.2 mM Na3VO4). Test compounds were reconstituted in 100% DMSO and added to the reaction to give a final DMSO concentration of 5%. Reactions were initiated by the addition 20 μl per well of kinase buffer containing 200-300 ng purified cytoplasmic domain KDR protein (BPS Bioscience, San Diego, Calif.). Following a 15 minute incubation at 30° C., the reactions were washed 2 mLs per well PBS-T. 100 μl of a monoclonal anti-phosphotyrosine antibody-peroxidase conjugate diluted 1:10,000 in PBS-T was added to the wells for 30 minutes. Following a 2 mLs per well wash with PBS-Tween-20, 100 μl of O-Phenylenediamine Dihydrochloride in phosphate-citrate buffer, containing urea hydrogen peroxide, was added to the wells for 7-10 minutes as a colorimetric substrate for the peroxidase. The reaction was terminated by the addition of 100 μl of 2.5N H2SO4 to each well and read using a microplate ELISA reader set at 492 nm.
Measurement: IC50 = 1470 nM (pChEMBL 5.829999923706055)
Descriptors: MW 441.49, ALogP 4.73, HBA 4, HBD 3, PSA 95.06, QED 0.55
Compound (SMILES): N=C(N)Nc1ccc(C(=O)Oc2ccc(CCC(=O)N(Cc3ccccc3)C(=O)O)c(Cl)c2)cc1
InChIKey: YCNDBVHYSLOGCW-UHFFFAOYSA-N
Target: Serine protease 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P07477
Assay: Inhibition Assay: The compound was dissolved in dimethyl sulfoxide (DMSO), and diluted to an arbitrary concentration (A). A was 100-fold diluted with a buffer (0.1 M Tris (pH 8.0), 0.15 M NaCl, 10 mM CaCl2, 0.05% Brij38) (B). The r-h trypsin was diluted with a buffer to 0.088 ug/mL, and the m-trypsin was diluted with a buffer to 1/50 (C). The dilution ratio of the m-trypsin (1/50) was set to exhibit the same activity as the 0.088 ug/mL r-h trypsin as determined by kinetic analysis. The substrate solution of a substrate for the enzyme reaction, BZiPAR, (Rhodamine Reference Substrate) was diluted with a buffer to 5 umol/L (D). B; 5 uL, C; 5 uL, and D; 10 uL were added to a 384-plate, and incubated at room temperature for 30 minutes. The fluorescent signals were detected with Ex/Em = 497/520 using Tecan Safire Fluorometer. The compound was reviewed from 2500 nM to its 3-fold value, 0.0075 nM, at 12 concentrations, and the inhibitory rate of each compound was calculated.
Measurement: IC50 = 0.51 nM (pChEMBL 9.289999961853027)
Descriptors: MW 494.94, ALogP 4.5, HBA 5, HBD 4, PSA 145.81, QED 0.16
Compound (SMILES): O=C(O)C12CCC(C(=O)N3CC[C@](c4ccc(C(F)(C(F)(F)F)C(F)(F)F)cc4)(S(=O)(=O)c4ccc(F)cc4)C3)(CC1)CC2
InChIKey: BFUUZNNTBJZLDG-WNMGUVTHSA-N
Target: Nuclear receptor ROR-gamma (SINGLE PROTEIN), Homo sapiens
UniProt: P51449
Assay: Scintillation Proximity Assay (SPA): The binding of potential ligands to RORγ is measured by competition with [3H] 25-hydroxycholesterol (Perkin Elmer NET674250UC) using a scintillation proximity assay (SPA) binding assay. The ligand binding domain of human RORγ (A262-S507) with an N-terminal His tag is expressed in E. coli and purified using nickel affinity chromatography. 15 ug/well RORγ (A262-S507) is incubated with test compound at varying concentrations in 3-fold serial dilution, with final concentrations ranging from 16.6 uM to 0.28 nM for 10 min at room temperature in PBS buffer (Invitrogen #14190-144) containing 0.5% fatty acid free BSA (Gemini Bio-Products, Cat. #700-107P) and 0.1% Glycerol (Sigma Cat#G5516). 10 nM of [3H] 25-hydroxycholesterol is then added, and the reaction is incubated for 10 min. 10 mg/mL of Copper-His Tag-PVT beads (Perkin Elmer cat #RPNQ0095) are added, and the mixture is incubated for 60 min. The reaction is read on a TopCount Microplate scintillation plate reader (Perkin Elmer). The competition data of the test compound over a range of concentrations was plotted as percentage inhibition of radioligand specifically bound in the absence of test compound (percent of total signal). After correcting for non-specific binding, IC50 values were determined. The IC50 value is defined as the concentration of test compound needed to reduce [3H] 25-hydroxycholesterol specific binding by 50% and is calculated using the four parameter logistic equation to fit the normalized data.
Measurement: IC50 = 60 nM (pChEMBL 7.21999979019165)
Descriptors: MW 653.59, ALogP 6.44, HBA 4, HBD 1, PSA 91.75, QED 0.29
Compound (SMILES): NC(=O)c1cc(N2CCNCC2C(N)=O)nc(-c2ccc(Oc3ccc(F)cc3)cc2)n1
InChIKey: HXLNLMXKKIOHJX-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: FLIPR Assay: In vitro assays were performed in a recombinant cell line expressing cDNA encoding the alpha subunit (Nav1.7, SCN9a, PN1, NE) of human Nav1.7 (Accession No. NM002977). The cell line was provided by investigators at Yale University (Cummins et al, J. Neurosci. 18(23): 9607-9619 (1998)). For dominant selection of the Nav1.7-expressing clones, the expression plasmid co-expressed the neomycin resistance gene. The cell line was constructed in the human embryonic kidney cell line, HEK293, under the influence of the CMV major late promoter, and stable clones were selected using limiting dilution cloning and antibiotic selection using the neomycin analogue, G418. Recombinant beta and gamma subunits were not introduced into this cell line. Additional cell lines expressing recombinant Nav1.7 cloned from other species can also be used, alone or in combination with various beta subunits, gamma subunits or chaperones.
Measurement: IC50 = 260 nM (pChEMBL 6.579999923706055)
Descriptors: MW 436.45, ALogP 1.44, HBA 7, HBD 3, PSA 136.46, QED 0.53
Compound (SMILES): COc1cc(OC)cc(N(C[C@H](OC(C)=O)C(F)(F)F)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: HHAMLGNIKIGSCW-DEOSSOPVSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 181.97 nM (pChEMBL 6.739999771118164)
Descriptors: MW 515.49, ALogP 4.68, HBA 9, HBD 0, PSA 91.6, QED 0.31
Compound (SMILES): N#CCNc1cc(-c2ccc(OCc3cccc(C(=O)N4CCC[C@H]4C(=O)O)c3)cc2)cc(F)c1F
InChIKey: XLMAQBUFOOVASV-DEOSSOPVSA-N
Target: Glycogen [starch] synthase, muscle (SINGLE PROTEIN), Homo sapiens
UniProt: P13807
Assay: Glycogen Synthase Activity Assay: To a polystyrene 96-well plate, a solution containing 30 mM glycylglycine (pH 7.3), 40 mM KCl, 20 mM MgCl2, 9.2% DMSO containing one of the test compounds at various concentrations, and 10 mM glucose-6-phosphate (Sigma-Aldrich Corporation, G7879) was added by 12 uL/well. Next, a substrate solution containing 30 mM glycylglycine (pH 7.3), 4.3 mg/mL of glycogen (Sigma-Aldrich Corporation, G8876), 21.6 mM UDP-glucose (Sigma-Aldrich Corporation, U4625), 21.6 mM phosphoenolpyruvic acid (Sigma-Aldrich Corporation, P0564), and 4.05 mM NADH (Sigma-Aldrich Corporation, N8129) was added by 18 uL/well. Further, an enzyme solution containing 50 mM Tris-HCl (pH 8.0), 27 mM DTT (Nacalai Tesque, Inc., 14128-04), 0.2 mg/mL of bovine serum albumin, 0.17 mg/mL of the glycogen synthase, 1.5 uL of a pyruvate kinase/lactate dehydrogenase solution (Sigma-Aldrich Corporation, P0294) was added by 18 uL/well to prepare a reaction solution. After the reaction solution was incubated (at 30° C. for 25 minutes for Examples 1 to 10, at 37° C. for 20 minutes for Examples 11 to 75), the absorbance at 340 nm was measured using Benchmark Plus (Bio-Rad Laboratories, Inc.).
Measurement: EC50 = 54 nM (pChEMBL 7.269999980926514)
Descriptors: MW 491.49, ALogP 4.84, HBA 5, HBD 2, PSA 102.66, QED 0.44
Compound (SMILES): O=C(CN1CCOCC1)Nc1cc(Cc2n[nH]c(=O)c3c2CCCC3)ccc1F
InChIKey: GZBWWLFDBUYPRZ-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P09874
Assay: PARP Enzyme Inhibition Assay: PARP1 assay was conducted in PARP assay buffer containing 50 mM Tris pH 8.0, 1 mM DTT, 4 mM MgCl2. PARP reactions contained 1.5 uM [3H]-NAD+ (1.6 uCi/mmol), 200 nM biotinylated histone H1, 200 nM sIDNA, and 1 nM PARP enzyme. Auto reactions utilizing SPA bead-based detection were carried out in 100 uL volumes in white 96 well plates. Reactions were initiated by adding 50 ul of 2xNAD+ substrate mixture to 50 uL of 2x enzyme mixture containing PARP and DNA. These reactions were terminated by the addition of 150 uL of 1.5 mM benzamide (~1000-fold over its IC50). 170 uL of the stopped reaction mixtures were transferred to streptavidin Flash Plates, incubated for 1 hour, and counted using a TopCount microplate scintillation counter.
Measurement: Ki = 19 nM (pChEMBL 7.71999979019165)
Descriptors: MW 400.45, ALogP 1.65, HBA 5, HBD 2, PSA 87.32, QED 0.8
Compound (SMILES): O=C(Nc1cnn(S(=O)(=O)c2cc(Cl)ccc2Cl)c1)c1ccccn1
InChIKey: PFNHJZNUOHOVJT-UHFFFAOYSA-N
Target: Metabotropic glutamate receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q14833
Assay: Glutamate Receptor Activity: The disclosed compounds and compositions can be evaluated for their ability to act as a potentiator of metabotropic glutamate receptor activity, in particular mGluR4 activity, by any suitable known methodology known in the art. For example, Chinese Hamster Ovary (CHO) cells transfected with human mGluR4 or HEK cells co-transfected with rat mGluR4 and the G-protein regulated Inwardly Rectifying Potassium channel (GIRK) were plated in clear bottom assay plates for assay in a Hamamatsu FDSS Fluorometric Plate Reader. The cells were loaded with either a Ca2+-sensitive fluorescent dye or the thallium responsive dye and the plates were washed and placed into a suitable kinetic plate reader. For human mGluR4 assays, a fluorescence baseline was established for 3-5 seconds, the disclosed compounds were then added to the cells, and the response in cells was measured.
Measurement: EC50 = 197 nM (pChEMBL 6.710000038146973)
Descriptors: MW 397.24, ALogP 3.07, HBA 6, HBD 1, PSA 93.95, QED 0.73
Compound (SMILES): CC1(C)Cc2nc(C3CCN(c4ncc(C#N)cn4)CC3)c([C@@H](F)c3ccc(C(F)(F)F)cc3)c(C3CCC(F)(F)CC3)c2[C@@H](O)C1
InChIKey: ANPNVRIFRYOAPY-YZNIXAGQSA-N
Target: Cholesteryl ester transfer protein (SINGLE PROTEIN), Homo sapiens
UniProt: P11597
Assay: CETP Inhibition Assay: A recombinant human CETP protein (manufactured by Roar Biomedical Inc.; 4.5 ng), the acceptor lipoprotein described in (2) above (32.5 ug) and 5,5'-dithio-bis-(2-nitrobenzoic acid) (7 mM, 15 ul) were taken in a 96 well plate and the total amount of the mixture was adjusted to 48.5 ul with the PBS solution. The test compound [DMSO solution (concentration: 0.15, 0.5, 1.5, 5, 15, 50, 150 and 500 uM; 1.5 ul] was added to each well and the mixture was incubated in a thermostatic bath at 37° C. for 60 minutes. The reconstituted HDL (50 ul) described in (1) above was added to each well and the mixture was reacted in a thermostatic bath at 37° C. for 60 minutes. The 96 well plate was moved onto ice and a precipitation reagent [a mixed solution of magnesium chloride (60 mM) and 0.1% dextran sulfate [1/1 (v/v)]; 15 ul] was added to each well, and then the mixture was allowed to stand on ice for 15 minutes.
Measurement: IC50 = 217 nM (pChEMBL 6.659999847412109)
Descriptors: MW 657.7, ALogP 8.5, HBA 6, HBD 1, PSA 85.93, QED 0.28
Compound (SMILES): CC(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@@H](CCC(N)=O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@H](C(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@H](C(=O)N[C@@H](Cc1ccccc1)C(=O)NCC(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(N)=O)C(C)C)C(C)C
InChIKey: RIMWGBJDGJAILI-AAYHSTQHSA-N
Target: Tissue factor pathway inhibitor (SINGLE PROTEIN), Homo sapiens
UniProt: P10646
Assay: Binding affinity to TFPI (unknown origin) incubated for 1.5 hrs using tracer peptide JBT0131 by screening ELISA
Measurement: IC50 = 679 nM (pChEMBL 6.170000076293945)
Descriptors: MW 2529.04
Compound (SMILES): CCOC(=O)N(C)C(=O)c1c(-c2ccc(F)cc2)oc2cc(N(C)S(C)(=O)=O)c(-c3ccc4c(n3)-c3cc5c(F)cccc5n3CO4)cc12
InChIKey: HNUBFPDEZQLPHD-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1a (isolate H) (HCV)
UniProt: P27958
Assay: ELISA-Based Assay: Measurement of inhibition by compounds was performed using the HCV replicon system. Several different replicons encoding different HCV genotypes or mutations were used. In addition, potency measurements were made using different formats of the replicon assay, including different ways of measurements and different plating formats. See Jan M. Vrolijk et al., A replicons-based bioassay for the measurement of interferons in patients with chronic hepatitis C, 110 J. VIROLOGICAL METHODS 201 (2003); Steven S. Carroll et al., Inhibition of Hepatitis C Virus RNA Replication by 2'-Modified Nucleoside Analogs, 278(14) J. BIOLOGICAL CHEMISTRY 11979 (2003). However, the underlying principles are common to all of these determinations, and are outlined below.Stable neomycin phosphotransferase encoding replicons-harboring cell lines were used, so all cell lines were maintained under G418 selection prior to the assay. Potency was determined using a cell ELISA assay with an antibody to the replicons encoded NS3/4a protease. See Caterina Trozzi et al., In Vitro Selection and Characterization of Hepatitis C Virus Serine Protease Variants Resistant to an Active-Site Peptide Inhibitor, 77(6) J. Virol. 3669 (2003).
Measurement: IC50 = 42 nM (pChEMBL 7.380000114440918)
Descriptors: MW 686.69, ALogP 7.04, HBA 9, HBD 0, PSA 124.18, QED 0.18
Compound (SMILES): CO[C@H]1CC[C@H](c2cnc(N)c(-c3ccc(C(=O)NCc4ccccc4)c(F)c3)n2)CC1
InChIKey: KYWPHHGZULMIRX-UAPYVXQJSA-N
Target: Mitogen-activated protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P28482
Assay: Kinase Assay (New): Compound potency against activated ERK2 is determined using a kinase assay that measures ERK2-catalyzed phosphorylation of biotinylated ERKtide peptide substrate ([Biotin]-AHA-K-R-E-L-V-E-P-L-T-P-S-G-E-A-P-N-Q-A-L-L-R-[NH2], the peptide sequence derived from EGF receptor: SEQ ID NO:1). The assay is carried out in 50 mM HEPES [pH 7.5], 5 mM MgCl2, 1 mM DTT, 0.01% Tween-20, 0.05% BSA using 0.25 nM ERK2, 200 nM ERKtide peptide and 35 uM ATP (all concentrations are final in the reaction) in a total volume of 10.25 uL. A 16-point, half-log dilution series of compounds at 41x final concentration is used for generating IC50 curves. Compound dilution series are prepared in 100% DMSO. ERK2 is preincubated with compounds for 30 minutes at ambient temperature. Reaction is initiated by addition of a substrate cocktail of ERKtide peptide and ATP and is allowed to proceed for 2-3 hours at ambient temperature. Reaction is terminated by addition of 10 uL of a 2x stop buffer consisting of 100 mM Tris-Cl [pH 7.5], 25 mM EDTA, 0.01% Tween 20, 10 ug/mL of AlphaScreen Protein A Acceptor Beads, 10 ug/mL of Streptavidin Donor Beads (PerkinElmer, Waltham, Mass.), and 1.4 ug/mL phospho-EGF Receptor (Thr669) antibody (Cat #3056, Cell Signaling Technology, Danvers, Mass.). Terminated reactions are read, after overnight incubation in the dark, on an EnVision Multilabel Plate Reader (PerkinElmer, Waltham, Mass.), with excitation and emission wavelengths set to 680 nm and 570 nm, respectively. IC50 values are determined using a four-parameter fit.
Measurement: IC50 = 18.9 nM (pChEMBL 7.71999979019165)
Descriptors: MW 434.52, ALogP 4.47, HBA 5, HBD 2, PSA 90.13, QED 0.6
Compound (SMILES): O=c1[nH]c(CN2CC(OCc3ccc(F)cc3)C2)nn2c(C3CCOCC3)ncc12
InChIKey: QEJJXMNBAFWGJK-UHFFFAOYSA-N
Target: High affinity cGMP-specific 3',5'-cyclic phosphodiesterase 9A (SINGLE PROTEIN), Homo sapiens
UniProt: O76083
Assay: Inhibition Assay: A PDE9 assay may for example, be performed as follows: The assay is performed in 60 uL samples containing a fixed amount of the relevant PDE enzyme (sufficient to convert 20-25% of the cyclic nucleotide substrate), a buffer (50 mM HEPES 7.6; 10 mM MgCl2; 0.02% Tween20), 0.1 mg/ml BSA, 225 pCi of 3H-labelled cyclic nucleotide substrate, tritium labeled cAMP to a final concentration of 5 nM and varying amounts of inhibitors. Reactions are initiated by addition of the cyclic nucleotide substrate, and reactions are allowed to proceed for one hr at room temperature before being terminated through mixing with 15 uL 8 mg/mL yttrium silicate SPA beads (Amersham). The beads are allowed to settle for one hr in the dark before the plates are counted in a Wallac 1450 Microbeta counter.
Measurement: IC50 = 10 nM (pChEMBL 8)
Descriptors: MW 413.45, ALogP 1.85, HBA 7, HBD 1, PSA 84.75, QED 0.66
Compound (SMILES): COC(=O)Nc1nc2ccc(C(=O)c3ccccc3C(=O)Nc3cc(C)ccc3F)cc2[nH]1
InChIKey: GCRJRFKBIWWHPI-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition of B-Raf (unknown origin) pre-incubated for 30 mins by luciferase-luciferin-coupled chemiluminescence assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 446.44, ALogP 4.67, HBA 5, HBD 3, PSA 113.18, QED 0.39
Compound (SMILES): Cn1cc(-c2cc(NS(C)(=O)=O)ccc2OCc2ccccc2)c2cc[nH]c2c1=O
InChIKey: ZXUXOEACJYQVPI-UHFFFAOYSA-N
Target: Bromodomain-containing protein 4 (SINGLE PROTEIN), Homo sapiens
UniProt: O60885
Assay: Bromodomain Domain Binding Assay: A time-resolved fluorescence resonance energy transfer (TR-FRET) assay was used to determine the affinities of compounds of the Examples listed in Table 1 for each bromodomain of human BRD4. His-tagged first (BD1: amino acids K57-E168) and second (BD2: amino acids E352-E168) bromodomains of human BRD4 were expressed and purified. An Alexa647-labeled BET-inhibitor was used as the fluorescent probe in the assay. Compound dilution series were prepared in DMSO via a 3-fold serial dilution from 2.5 mM to 42 nM. Compounds were then diluted 6:100 in assay buffer (20 mM Sodium Phosphate, pH 6.0, 50 mM NaCl, 1 mM Ethylenediaminetetraacetic acid disodium salt dihydrate, 0.01% Triton X-100, 1 mM DL-Dithiothreitol) to yield 3× working solutions. Six microliters (L) of the working solution was then transferred to white, low-volume assay plates (Costar #3673). A 1.5× assay mixture containing His-tagged bromodomain, Europium-conjugated anti-His antibody (Invitrogen PV5596) and the Alexa-647-conjugated probe molecule was also prepared. Twelve L of this solution were added to the assay plate to reach a final volume of 18 L. The final concentration of 1× assay buffer contains 2% DMSO, 50 M-0.85 nM compound, 8 nM His-tagged bromodomain, 1 nM Europium-conjugated anti-His-tag antibody and 100 nM or 30 nM probe (for BDI or BDII, respectively). After a one-hour incubation at room temperature, TR-FRET ratios were determined using an Envision multilabel plate reader (Ex 340, Em 495/520).
Measurement: Ki = 18 nM (pChEMBL 7.75)
Descriptors: MW 423.49, ALogP 3.48, HBA 5, HBD 2, PSA 93.19, QED 0.5
Compound (SMILES): Cc1ccc(-n2nc(C3CCN([S+]([O-])Cc4noc5ccccc45)CC3)cc2NC(=O)c2cnn3cccnc23)cc1
InChIKey: ONVHOXOVXYATQL-UHFFFAOYSA-N
Target: Interleukin-1 receptor-associated kinase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NWZ3
Assay: Kinase Assay: The kinase activity of IRAK-4 is determined by its ability to catalyze the phosphorylation of a fluorescent polypeptide substrate. The extent of phosphorylation is measured using the IMAP technology (Molecular Devices) where the phosphorylated fluorescent substrate binds to the large M(III)-based nanoparticles which reduces the rotational speed of the substrate and thus increases its fluorescent polarization (FP). 20 μl reaction mixture contains 10 mM TriHCl, pH 7.2, 0.5 nM GST tagged IRAK-4 (SignalChem), 100 nM fluorescent peptide substrate (RP7030, Molecular Devices), 100 μM ATP, 1 mM DTT, 1 mM MgCl2, and 0.01% Tween 20. The reaction is initiated by the addition of ATP. After incubation for 30 minutes at 25° C., 60 μl of Progressive IMAP Reagent (Molecular Devices) is added to stop the reaction. Change in RP7030's FP is determined by a FP reader (Analyst HT, LH, BioSystems).
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 580.67, ALogP 4.66, HBA 10, HBD 1, PSA 129.44, QED 0.27
Compound (SMILES): COc1cc2c(Nc3ccc(Br)c(Cl)c3)ncnc2cc1O[C@@H]1C[C@H]2OCC3(OCCO3)[C@H]2O1
InChIKey: XCAOLOJNZVNRSK-HKBOAZHASA-N
Target: Ephrin type-B receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P54760
Assay: Inhibition of recombinant human N-terminal His6-tagged EphB4 kinase domain (E605 to E890 residues) expressed in baculovirus using poly-AEKY as substrate after 150 mins in presence of gamma33P-ATP by liquid scintillation spectrometry
Measurement: IC50 = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 550.79, ALogP 4.43, HBA 9, HBD 1, PSA 93.19, QED 0.5
Compound (SMILES): CC(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(=O)N[C@H](C(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@H](C(=O)N[C@@H](CC(=O)O)C(=O)NCC(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@H](C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(N)=O)[C@@H](C)O)C(C)C)C(C)C
InChIKey: AOKUTFOEHXSLTL-QBVKEVBNSA-N
Target: Tissue factor pathway inhibitor (SINGLE PROTEIN), Homo sapiens
UniProt: P10646
Assay: Binding affinity to TFPI (unknown origin) incubated for 1.5 hrs using tracer peptide JBT0131 by screening ELISA
Measurement: IC50 = 250 nM (pChEMBL 6.599999904632568)
Descriptors: MW 2369.72
Compound (SMILES): N#Cc1ccc(Cl)cc1-c1ccn(C(Cc2ccccc2)C(=O)Nc2ccc(-c3nnn[nH]3)cc2)c(=O)c1
InChIKey: QLAFRGZBWLGFNO-UHFFFAOYSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: Inhibition Assay: Test substances are dissolved in dimethyl sulphoxide and serially diluted in dimethyl sulphoxide (3000 μM to 0.0078 μM; resulting final concentrations in the test: 50 μM to 0.00013 μM). In each case 1 μl of the diluted substance solutions are placed into the wells of white microtitre plates from Greiner (384 wells). 20 μl of assay buffer (50 mM of Tris/HCl pH 7.4; 100 mM of sodium chloride solution; 5 mM of calcium chloride solution; 0.1% of bovine serum albumin) and 20 μl of factor XIa from Kordia (0.45 nM in assay buffer) are then added successively. After 15 min of incubation, the enzyme reaction is started by addition of 20 μl of the factor XIa substrate Boc-Glu(OBzl)-Ala-Arg-AMC dissolved in assay buffer (10 μM in assay buffer) from Bachem, the mixture is incubated at room temperature (22° C.) for 30 min and fluorescence is then measured (excitation: 360 nm, emission: 460 nm).
Measurement: IC50 = 7.6 nM (pChEMBL 8.119999885559082)
Descriptors: MW 521.97, ALogP 4.64, HBA 7, HBD 2, PSA 129.35, QED 0.32
Compound (SMILES): CCC1CC(=O)NN=C1c1ccc2nc(-c3ccc(OCCN4CCSCC4)cc3)oc2c1
InChIKey: PZLOAEHOORDZAA-UHFFFAOYSA-N
Target: Glucose-dependent insulinotropic receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q8TDV5
Assay: AlphaScreen cAMP Assay: MIN6 cells [Miyazaki J et al. Endocrinology. 1990 July; 127(1):126-32] were stably transfected with an expression vector for human GPR119 cDNA (Acc. No. NP-848566). Min-6/hGPR119 cells are cultured in DMEM, 10% FBS, 50 uM beta-mercaptoethanol, 0.3 mg/mL Geniticin, 2 mM GlutaMAX at 37 C. 5% CO2. For the assay, the cells are placed in Optiplates (white, 384-well, 160W-barcoded, TC, sterile with lid, Cat. No. #6007688 (Perkin Elmer); 10000 cells/well; 50 ul). The plates covered with lids are then incubated for 24 hours at 37 C./5% CO2.After the medium has been sucked out of the wells completely, 10 ul of the test compound are added, the compounds are diluted beforehand with stimulating buffer (140 mM NaCl, 3.6 mM KCl, 0.5 mM NaH2PO4, 0.5 mM MgSO4, 1.5 mM CaCl2, 10 mM Hepes, 5 mM NaHCO3; pH 7.4. 0.5 mM IBMX and 0.1% BSA, the final DMSO concentration is 1%). After 45 minutes' incubation at RT, the cAMP concentrations are determined using the AlphaScreen cAMP Assay.
Measurement: EC50 = 62 nM (pChEMBL 7.210000038146973)
Descriptors: MW 464.59, ALogP 4.17, HBA 7, HBD 1, PSA 79.96, QED 0.57
Compound (SMILES): COc1ccc(C(C)C)cc1C1=C(CN(Cc2cc(C(F)(F)F)cc(C(F)(F)F)c2)c2ncc(-c3cnn(C)c3)cn2)CCc2ccccc21
InChIKey: SYPLOXXEJRTVHJ-UHFFFAOYSA-N
Target: Cholesteryl ester transfer protein (SINGLE PROTEIN), Homo sapiens
UniProt: P11597
Assay: Inhibition of recombinant CETP (unknown origin) assessed as inhibition of BODIPY-labeled cholesteryl transfer to fluorescent substrate by fluorimetric assay
Measurement: IC50 = 220 nM (pChEMBL 6.659999847412109)
Descriptors: MW 691.72, ALogP 9.5, HBA 6, HBD 0, PSA 56.07, QED 0.14
Compound (SMILES): COC(=O)[C@@H](C)CN(c1sc2ccccc2c1C)S(=O)(=O)c1ccccc1
InChIKey: WREWEOWZYOZQJQ-AWEZNQCLSA-N
Target: Transient receptor potential M8 protein (SINGLE PROTEIN), Canis lupus familiaris
UniProt: Q1A7N1
Assay: Functional Assay: The functional activity of compounds of the formula (I) was determined by measuring changes in intracellular calcium concentration using a Ca2+-sensitive fluorescent dye. The changes in fluorescent signal were monitored by a fluorescence plate reader, either a FLIPR (Molecular Devices) or FDSS (Hamamatsu). Increases in intracellular Ca2+ concentration were readily detected upon activation with icilin.At 24 hrs prior to assay, HEK293 cells stably expressing canine TRPM8 were seeded in culture medium in black wall, clear-base poly-D-lysine coated 384-well plates (BD Biosciences, NJ, USA) and grown overnight in 5% CO2 at 37° C. On assay day, growth media was removed and cells were loaded with Calcium 3 Dye (Molecular Devices) for 35 min at 37° C., under 5% CO2 and then for 25 min at room temperature and atmosphere. Subsequently, cells were tested for agonist-induced increases in intracellular Ca2+ levels using FLIPR or FDSS. Cells were challenged with a compound of the Formula (I) (at varying concentrations) and intracellular Ca2+ was measured for 5 min prior to the addition of icilin to all wells to achieve a final concentration that produces approximately an 80% maximal response. EC50 or IC50 values for compounds of the present invention were determined from eight-point dose-response studies.
Measurement: IC50 = 18 nM (pChEMBL 7.75)
Descriptors: MW 403.53, ALogP 4.21, HBA 5, HBD 0, PSA 63.68, QED 0.58
Compound (SMILES): O=C1NCc2ccc(-c3cccc(OCC(O)CN4CCc5ccccc5C4)c3)cc21
InChIKey: OZLUBKSIOHYRGU-UHFFFAOYSA-N
Target: PRMT5/MEP50 complex (PROTEIN COMPLEX), Homo sapiens
UniProt: Q9BQA1
Assay: Inhibition of recombinant human fetal brain FLAG-tagged PRMT5/full length human testis 6His-tagged MEP50 expressed in baculovirus infected insect Sf9 cells using human histone H4 (1 to 15 residues) as substrate preincubated for 30 mins followed by [3H]-SAM addition
Measurement: IC50 = 101 nM (pChEMBL 7)
Descriptors: MW 414.51, ALogP 3.4, HBA 4, HBD 2, PSA 61.8, QED 0.65
Compound (SMILES): COc1cc2c(cc1OC)C(=O)/C(=C/c1ccc(N3CCN(C(=O)c4ccccc4)CC3)cc1)C2
InChIKey: GJSRQFLUIYWOGE-XQNSMLJCSA-N
Target: Acetylcholinesterase (SINGLE PROTEIN), Electrophorus electricus
UniProt: O42275
Assay: Inhibition of Electric eel AChE using acetylthiocholine iodide as substrate preincubated for 5 mins followed by substrate addition measured after 2 mins by Ellman's method
Measurement: IC50 = 1701 nM (pChEMBL 5.769999980926514)
Descriptors: MW 468.55, ALogP 4.49, HBA 5, HBD 0, PSA 59.08, QED 0.52
Compound (SMILES): Cc1c(Cl)cc(O)c2nc(CCc3nc(N4CCCC4)nn3C)nn12
InChIKey: HGXGMFATLSNPBE-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y233
Assay: Scintillation Proximity Assay: The human PDE10A full length assay was performed in 96-well micro titer plates. The reaction mixture of 50 μl contained 20 mM HEPES pH=7.5/10 mM MgCl2/0.05 mg/ml BSA (Sigma cat. # A-7906), 50 nM cGMP (Sigma, cat. # G6129) and 50 nM [3H]-cGMP (GE Healthcare, cat. # TRK392 S.A. 13.2 Ci/mmol), 3.75 ng/well PDE10A enzyme (Enzo Life Science, Lausen, Switzerland cat # SE-534) with or without a specific test compound. A range of concentrations of the potential inhibitor was used to generate data for calculating the concentration of inhibitor resulting in 50% of the effect (e.g. IC50, the concentration of the competitor inhibiting PDE10A activity by 50%). Non-specific activity was tested without the enzyme. The reaction was initiated by the addition of the substrate solution (cGMP and [3H]-cGMP) and allowed to progress for 20 minutes at room temperature. The reaction was terminated by adding 25 μl of YSi-SPA scintillation beads (GE Healthcare, cat. # RPNQ0150) in 18 mM zinc sulphate solution (stop reagent). After 1 h under shaking, the plate was centrifuged one minute at 170 g to allow beads to settle. Afterwards, radioactive counts were measured on a Perkin Elmer Top-Count Scintillation plate reader.
Measurement: IC50 = 4.22 nM (pChEMBL 8.380000114440918)
Descriptors: MW 361.84, ALogP 1.91, HBA 8, HBD 1, PSA 84.37, QED 0.76
Compound (SMILES): COc1ccc(C2(C(F)(F)F)CC(c3ccc(C)cc3)=C(C#N)C(=O)N2)cc1
InChIKey: JWJSLVALDGJEQA-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Mus musculus
UniProt: Q921V5
Assay: Inhibition of mouse recombinant MGAT2 expressed in Sf9 cell membrane using 2-oleoylglycerol and oleoyl-CoA as substrates incubated for 10 mins by LCMS assay
Measurement: IC50 > 1271 nM (pChEMBL 5.895999908447266)
Descriptors: MW 386.37, ALogP 4.26, HBA 3, HBD 1, PSA 62.12, QED 0.86
Compound (SMILES): [C-]#[N+]c1cnc2c(C(=O)Nc3cc(C4CCN(S(C)(=O)=O)CC4)nn3-c3ccc(C)cn3)cnn2c1
InChIKey: BRSACOABJUCYBG-UHFFFAOYSA-N
Target: Interleukin-1 receptor-associated kinase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NWZ3
Assay: Kinase Assay: The kinase activity of IRAK-4 is determined by its ability to catalyze the phosphorylation of a fluorescent polypeptide substrate. The extent of phosphorylation is measured using the IMAP technology (Molecular Devices) where the phosphorylated fluorescent substrate binds to the large M(III)-based nanoparticles which reduces the rotational speed of the substrate and thus increases its fluorescent polarization (FP). 20 μl reaction mixture contains 10 mM TriHCl, pH 7.2, 0.5 nM GST tagged IRAK-4 (SignalChem), 100 nM fluorescent peptide substrate (RP7030, Molecular Devices), 100 μM ATP, 1 mM DTT, 1 mM MgCl2, and 0.01% Tween 20. The reaction is initiated by the addition of ATP. After incubation for 30 minutes at 25° C., 60 μl of Progressive IMAP Reagent (Molecular Devices) is added to stop the reaction. Change in RP7030's FP is determined by a FP reader (Analyst HT, LH, BioSystems).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 505.56, ALogP 2.56, HBA 9, HBD 1, PSA 131.74, QED 0.41
Compound (SMILES): CCOC(C(=O)c1ccc(-c2cc(OC)c(Cl)c(OC)c2)o1)c1ccc(-c2nnc(C)s2)cc1
InChIKey: QQHJRLKSCSRMEM-UHFFFAOYSA-N
Target: Dual specificity calcium/calmodulin-dependent 3',5'-cyclic nucleotide phosphodiesterase 1A (SINGLE PROTEIN), Homo sapiens
UniProt: P54750
Assay: Inhibtion Assay: The PDE assay medium consisted of (final concentrations); 50 mM Tris-HCL, pH 7.5; 8.3 mM MgCl2; 1.7 mM EGTA; 0.5 mg/mL bovine serum albumin; and substrate (H-cAMP or H-cGMP). The BSA was lyophilized powder, essentially fatty acid free, >=96% pure from Sigma (A6003). 89 ul of assay medium and 1 ul of Compound 1 in 100% DMSO were added to a 96-well SPA plate and incubated for 1 minute at 30° C. The assay was initiated by addition of 10 ul of PDE10, and incubated for 21 minutes. Then 50 ul of SPA beads were added and the plate was sealed, shaken, and incubated at room temperature for 1 hour. The plate was then counted in a Wallac 1450 Microbeta Trilux plate scintillation counter. For the PDE10 IC50 experiment, the substrate was 125 nM H-cGMP. For the selectivity experiments, the substrates were 37 nM H-cAMP for PDEs 3, 4, 7, and 8; and 37 nM H-cGMP for PDEs 1, 2, 5, 9, 10, and 11. In all cases substrate concentrations were below the KM. The consumption of substrate was less than 6%, indicating that substrate concentrations did not change appreciably during the assay.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 498.99, ALogP 6.4, HBA 8, HBD 0, PSA 83.68, QED 0.25
Compound (SMILES): NC(=O)c1ccc2[nH]c(-c3ccc(OC[C@H]4CC[C@H](O)CC4)cc3)nc2c1
InChIKey: LYTQPKFVJFPASC-SAZUREKKSA-N
Target: Serine/threonine-protein kinase Chk2 (SINGLE PROTEIN), Homo sapiens
UniProt: O96017
Assay: Inhibition of human CDS1 using N-terminal biotin labeled SGLYRSPSMPENLNRPR as substrate in presence of [gamma-33P]-ATP after 3 hrs by scintillation counting method
Measurement: IC50 = 400 nM (pChEMBL 6.400000095367432)
Descriptors: MW 365.43, ALogP 3.26, HBA 4, HBD 3, PSA 101.23, QED 0.65