lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CC[C@@H](CN1CCOCC1)N1C(=O)[C@@](C)(Cc2noc(=O)[nH]2)C[C@H](c2cccc(Cl)c2)[C@H]1c1ccc(Cl)cc1
InChIKey: UZLWVHOSWNFONP-NANZAVOOSA-N
Target: E3 ubiquitin-protein ligase Mdm2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00987
Assay: Homogenous Time-Resolved Fluorescence Assay (HTRF2 Assay): The standard assay conditions for the in vitro HTRF assay consisted of a 50 ul total reaction volume in black 384-well Costar polypropylene plates in 1×PBS buffer pH 7.4, 1 mM DTT, 0.1% BSA, 2.5 nM GST-hMDM2 (aa 1-188), 5 nM biotinylated-p53 (aa 1-83), 1.8 nM SA-XLent (Cisbio; Bedford, Mass.), 0.6 nM anti-GST cryptate monoclonal antibody (Cisbio; Bedford, Mass.) and 200 mM KF. Amino acid residues 1-188 of human MDM2 were expressed as an amino-terminal glutathione-S-transferase (GST) fusion protein (GST-hMDM2) in Escherichia coli. Residues 1-83 of human p53 were expressed as an amino-terminal AviTag™-TrxA-6×His fusion protein (biotinylated p53) in E. coli. Each protein was purified from cell paste by affinity chromatography.Specifically, 10 uL of GST-hMDM2 was incubated with 10 ul of diluted compound (various concentrations, serially diluted) in 10% DMSO for 20 minutes at room temperature. 20 uL of biotinylated-p53 was added to the GST-hMDM2+compound mixture, and then incubated at room temperature for 60 min. 10 uL of detection buffer consisting of SA-XLent, anti-GST cryptate antibody and KF was added to GST-hMDM2, biotinylated-p53 and compound reaction and left at room temperature to reach equilibrium for >4 hrs. The final concentration of DMSO in the reaction was 2%. Time-resolved fluorescence readings were measured on a microplate multilabel reader. Percentage of inhibition was calculated relative to nutlin-3. All assay conditions remained the same as described above, with the exception of the following changes in reagent concentrations: 0.2 nM GST-hMDM2 (1-188), 0.5 nM biotinylated-p53 (1-83), 0.18 nM SA-XLent, and 100 mM KF.
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 573.52, ALogP 5.09, HBA 6, HBD 1, PSA 91.67, QED 0.4
Compound (SMILES): COc1ccccc1Oc1ccc(C(=O)N(Cc2ccc(C(=O)O)cc2)CC(F)F)cc1
InChIKey: YOVTYVMJDJNYKX-UHFFFAOYSA-N
Target: Lysophosphatidic acid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q92633
Assay: Receptor Assay: Engagement of the LPA1 receptor by its ligand, oleoyl-L-Î±-lysophosphatidic acid (LPA), leads to the release of intracellular stores of calcium into the cytoplasm mediated through a Gq-driven increase in inositol triphosphate (IP3) levels. Gq is a heterotrimeric G protein subunit that activates phospholipase C. The ability of test compounds to prevent LPA-driven intracellular release of stored calcium from RH7777 cells expressing human LPA1 receptors was assayed by stimulating the cells with LPA in the presence of various test compounds to measure the test compounds' antagonism, i.e. activity in vitro.Â  Approximately 10,000 cells in normal culture medium (Dulbecco's Modified Eagle Media, 10% Foetal Calf Serum) were dispensed per assay well in a black clear bottom collagen coated 384 well plate (Beckton Dickinson). Cells were allowed to settle for thirty minutes at room temperature before being incubated overnight at 37° C., 5% CO2. Sixteen to twenty hours after dispensing, the cells were loaded with a calcium-sensitive fluorescent dye by replacing the culture medium with assay buffer (1x Hanks buffered saline, 25 mM HEPES, 0.1% w/v fatty acid free BSA (bovine serum albumin), pH7.4) containing 1.25 mM probenecid and 1x Calcium 5 Reagent (Molecular Devices). Cells were incubated at 37° C. for one hour to allow for dye uptake.
Measurement: IC50 = 34 nM (pChEMBL 7.46999979019165)
Descriptors: MW 441.43, ALogP 5.09, HBA 4, HBD 1, PSA 76.07, QED 0.5
Compound (SMILES): CCn1cc(-c2cc3c(cn2)cnn3-c2cccc(N3CCC[C@H](N)C3)n2)cn1
InChIKey: KSUJTDNWWVQOFC-KRWDZBQOSA-N
Target: Serine/threonine-protein kinase pim-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P1W9
Assay: Kinase Binding Assay: PIM-1, -2, and -3 enzymes were generated as fusion proteins expressed in bacteria and purified by IMAC column chromatography (Sun, X., Chiu, J. F., and He, Q. Y. (2005) Expert Rev. Proteomics, 2:649-657). A fluorescent-labeled Pim-specific peptide substrate, was custom synthesized by American Peptide Company (Sunnyvale, Calif.). Reaction Buffer contained 10 mM HEPES, pH 7.2, 10 mM MgCl2, 0.01% Tween 20, 2 mM DTT. Termination Buffer contained 190 mM HEPES, pH 7.2, 0.015% Brij-35, 0.2% Coating Reagent 3 (Caliper Life Sciences, Hopkinton, Mass.), 20 mM EDTA. Separation Buffer contained 100 mM HEPES, pH 7.2, 0.015% Brij-35, 0.1% Coating Reagent 3, 1:200 Coating Reagent 8 (Caliper Life Sciences, Hopkinton, Mass.), 10 mM EDTA and 5% DMSO.PIM reactions were carried out in a final volume of 10 uL per well in a 384-well plate. A standard enzymatic reaction, initiated by the addition of 5 uL 2Ã¿Â¿ATP and test compound to 5 uL of 2x enzyme and FAM-peptide, contained 20 uM PIM1, 50 uM PIM2, or 55 uM PIM3, 1 uM FAM-peptide, and 10 uM ATP, in Reaction Buffer. After 90 minutes of incubation at room temperature, the phosphorylation reaction was stopped by the addition of 10 uL Termination Buffer. The product and substrate in each independent reaction were separated on a 12-sipper microfluidic chip (Caliper Life Sciences, Hopkinton, Mass.) run on a Caliper LC3000 (Caliper Life Sciences, Hopkinton, Mass.). The separation of product and substrate was optimized by choosing voltages and pressure using Caliper's Optimizer software (Hopkinton, Mass.). The separation conditions used a downstream voltage of -500V, an upstream voltage of -2150V, and a screening pressure of -1.2 psi. The product and substrate fluorophore were excited at 488 nm and detected at 530 nm. Substrate conversion was calculated from the electropherogram using HTS Well Analyzer software (Caliper Life Sciences, Hopkinton, Mass.). Ki values for the test compound were calculated.
Measurement: Ki = 4.56 nM (pChEMBL 8.34000015258789)
Descriptors: MW 388.48, ALogP 2.63, HBA 8, HBD 1, PSA 90.68, QED 0.58
Compound (SMILES): CNC(C)C(=O)Nc1cc(-c2ncnc3c(C)nn(C)c23)cc(C#Cc2cc(F)c3ncccc3c2)n1
InChIKey: VKKPWLVOASHEOF-UHFFFAOYSA-N
Target: Baculoviral IAP repeat-containing protein 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13490
Assay: XIAP BIR3 & cIAP1 BIR3 Binding Assays (DELFIA): BIR3 domains (10 nM) were incubated with SMAC peptide (10 nM) in assay buffer (50 mM Tris, 120 mM NaCl, 0.1% BSA, 1 mM DTT, 0.05% TritonX100) for one hour at room temperature in the presence of inhibitory compounds. The assay mixture was transferred to a strepatvidin coated plate and incubated for one hour at room temperature to allow binding of the biotinylated peptide and associated BIR3 domains to the plate. After several washing steps Eu labeled anti-GST antibody (e.g. Perkin Elmer DELFIA Eu-N1-antiGST AD0250) was added to detect BIR3 domain-SMAC peptide interactions according to Perkin Elmer's instructions. Briefly, the antibody was added (dilution 1:5000 in Perkin Elmer DELFIA Assay Buffer 2013-01) and incubated for one hour. After 3 washing steps using Delfia Washing Buffer (Perkin Elmer DELFIA Wash 2013-05), Enhancement Solution (Perkin Elmer Enhancement Asolution 2013-02) was added and incubation continued for 10 minutes. Time resolved Europium fluorescence was measured in a Wallac Victor using Standard assay settings.
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 494.53, ALogP 3.37, HBA 8, HBD 2, PSA 110.51, QED 0.37
Compound (SMILES): Cc1nnc(C)n1-c1ccc(O[C@H]2c3cc(Cl)ccc3C[C@@H]2N2CC[C@@H](O)C2)c(Cl)c1
InChIKey: PXZHZDNVYZAURZ-JZWVFAODSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P48764
Assay: Inhibition of human NHE3 expressed in LAP1 cell assessed as intracellular pH recovery measured for 2 mins by CECF-AM dye based FLIPR assay
Measurement: IC50 = 90 nM (pChEMBL 7.050000190734863)
Descriptors: MW 459.38, ALogP 4.3, HBA 6, HBD 1, PSA 63.41, QED 0.63
Compound (SMILES): COC(=O)CN[C@H](CC1CCCCC1)C(=O)N[C@@H](Cc1ccccc1)C(=O)NCc1ccc(CN)cc1
InChIKey: MLHZEVRUXMVMRU-FTJBHMTQSA-N
Target: Plasma kallikrein (SINGLE PROTEIN), Homo sapiens
UniProt: P03952
Assay: In Vitro Inhibition Assay: Plasma kallikrein inhibitory activity in vitro was determined using standard published methods (see e.g. Johansen et al., Int. J. Tiss. Reac. 1986, 8, 185; Shori et al., Biochem. Pharmacol., 1992, 43, 1209; Stiirzebecher et al., Biol. Chem. Hoppe-Seyler, 1992, 373, 1025). Human plasma kallikrein (Protogen) was incubated at 37° C. with the fluorogenic substrate H-DPro-Phe-Arg-AFC and various concentrations of the test compound. Residual enzyme activity (initial rate of reaction) was determined by measuring the change in optical absorbance at 410 nm and the IC50 value for the test compound was determined.
Measurement: IC50 = 2100 nM (pChEMBL 5.679999828338623)
Descriptors: MW 508.66, ALogP 2.59, HBA 6, HBD 4, PSA 122.55, QED 0.31
Compound (SMILES): CCCOc1ccc(CCc2nc3cc(-c4c(C)noc4C)ccc3n2CCN2CCOCC2)cc1
InChIKey: CGWBJJZOKGZCSJ-UHFFFAOYSA-N
Target: Bromodomain-containing protein 4 (SINGLE PROTEIN), Homo sapiens
UniProt: O60885
Assay: Displacement of biotinylated ligand from BRD4 bromodomain-1 (unknown origin) by TR-FRET assay
Measurement: IC50 = 2900 nM (pChEMBL 5.539999961853027)
Descriptors: MW 488.63, ALogP 5.21, HBA 7, HBD 0, PSA 65.55, QED 0.31
Compound (SMILES): C[C@H](N)C/C=C\c1ccccc1
InChIKey: LLHJXESMOGGFAI-BSKOKIOFSA-N
Target: Sodium-dependent serotonin transporter (SINGLE PROTEIN), Rattus norvegicus
UniProt: P31652
Assay: Activity at SERT in rat synaptosomes obtained from whole brain minus striatum and cerebellum assessed as release of [3H]5-HT by liquid scintillation counting method
Measurement: EC50 = 663 nM (pChEMBL 6.179999828338623)
Descriptors: MW 161.25, ALogP 2.44, HBA 1, HBD 1, PSA 26.02, QED 0.72
Compound (SMILES): Cn1cc(-c2ccc3nc(NC(=O)c4cncc(-c5ccccc5)c4)cn3n2)c(-c2ccc(F)cc2)n1
InChIKey: FJBUTRXXIQJMHS-UHFFFAOYSA-N
Target: Casein kinase I isoform delta (SINGLE PROTEIN), Homo sapiens
UniProt: P48730
Assay: Kinase Assay: The kinase assay was performed in V-bottom 384-well plates. The final assay volume was 30 ul prepared from 15 ul additions of enzyme, substrates (fluoresceinated peptide FL-AHA-KRRRAL-PSER-VASLPGL-OH and ATP) and test compounds in assay buffer (20 mM HEPES pH 7.4, 30 mM MgCl2, 0.015% Brij35 and 4 mM DTT). The reaction was incubated at room temperature for 22 hours and terminated by adding 45 ul of 35 mM EDTA to each sample. The reaction mixture was analyzed on the Caliper LABCHIPÂ®3000 (Caliper, Hopkinton, Mass.) by electrophoretic separation of the unphosphorylated substrate and phosphorylated product Inhibition data were calculated by comparison of the no enzyme control reactions for 100% inhibition and vehicle-only reactions for 0% inhibition. The final concentration of reagents in the assay were 200 pM CK1ε or CK1δ, 50 uM ATP, 1.5 uM FL-AHA-KRRRAL-PSER-VASLPGL-OH, and 1.6% DMSO. Dose response curves were generated to determine the concentration required to inhibit 50% of the kinase activity (IC50). Compounds were dissolved at 10 mM in dimethylsulfoxide (DMSO) and evaluated at eleven concentrations. IC50 values were derived by non-linear regression analysis.
Measurement: IC50 = 0.5 nM (pChEMBL 9.300000190734863)
Descriptors: MW 489.51, ALogP 5.25, HBA 7, HBD 1, PSA 90, QED 0.36
Compound (SMILES): COc1ccncc1N1CCOc2cc(S(=O)(=O)Nc3nccs3)ccc21
InChIKey: IJASFADUIAWVTI-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: In Vitro PX Assay: HEK 293 cells stably transfected with human Nav1.7 were recorded in whole cell voltage clamp mode with the PatchXpress automated electrophysiology system (Molecular Devices, LLC, Sunnyvale, Calif.). Compound effects were measured on a partially inactivated state of the sodium channel. Cells were clamped to a holding potential yielding 20 to 50% inactivation. To elicit sodium current, channels were activated by pulsing to −10 mV for 20 msec. This voltage protocol was repeated at a rate of 0.1 Hz throughout the experiment. A single concentration of test compound was applied to cells for a duration of 3 minutes.
Measurement: IC50 = 5170 nM (pChEMBL 5.289999961853027)
Descriptors: MW 404.47, ALogP 2.88, HBA 8, HBD 1, PSA 93.65, QED 0.7
Compound (SMILES): O=C1N[C@H](c2cncc(C#CC3CC(F)(F)C3)c2)[C@@H](c2cccc(Cl)c2)O1
InChIKey: BVVFBBXWFCHQKF-QZTJIDSGSA-N
Target: Metabotropic glutamate receptor 5 (SINGLE PROTEIN), Homo sapiens
UniProt: P41594
Assay: Positive allosteric modulation activity at human mGluR5A expressed in HEK293(ZF) cells co-expressing rat glutamate-aspartate transporter assessed as increase in L-glutamate-induced Ca2+ flux preincubated for 60 mins followed by L-glutamate addition measured for 100 sec by calcium-4 dye based FLIPR assay
Measurement: EC50 = 66 nM (pChEMBL 7.179999828338623)
Descriptors: MW 388.8, ALogP 4.65, HBA 3, HBD 1, PSA 51.22, QED 0.76
Compound (SMILES): C=CC(=O)Nc1cccc(-n2c(=O)[nH]c3cnc(Nc4ccc(N5CCN(CC)CC5)cc4)nc32)c1
InChIKey: DAGUTUKQMPCUDA-UHFFFAOYSA-N
Target: Tyrosine-protein kinase BTK (SINGLE PROTEIN), Homo sapiens
UniProt: Q06187
Assay: Inhibition of recombinant human N-terminal His-tagged BTK expressed in baculovirus infected sf9 cells using poly(4:1 Glu,Tyr) as substrate by ADP-Glo kinase assay
Measurement: IC50 > 100 nM (pChEMBL 7)
Descriptors: MW 484.56, ALogP 3.12, HBA 8, HBD 3, PSA 111.18, QED 0.35
Compound (SMILES): CC(C)(O)c1nc(-c2cccc(CNC(=O)c3cnc(-c4ccccn4)nc3)c2)no1
InChIKey: VJCLAPUACUQZOV-UHFFFAOYSA-N
Target: Hematopoietic prostaglandin D synthase (SINGLE PROTEIN), Homo sapiens
UniProt: O60760
Assay: Inhibition of HPGDS (unknown origin) by fluorescence polarization assay
Measurement: IC50 = 7.48 nM (pChEMBL 8.130000114440918)
Descriptors: MW 416.44, ALogP 2.75, HBA 8, HBD 2, PSA 126.92, QED 0.49
Compound (SMILES): N#Cc1ccc(N2CCC(c3nnn4cnc5[nH]ccc5c34)CC2)cc1
InChIKey: MUUMCNOIQREEJX-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 190 nM (pChEMBL 6.71999979019165)
Descriptors: MW 343.39, ALogP 2.86, HBA 6, HBD 1, PSA 85.9, QED 0.6
Compound (SMILES): COc1ccc([C@@H]2CN(C)C[C@H]2NC(=O)c2ccc3cnc(Nc4cc(C)nn4C)cc3c2)cc1
InChIKey: UXYSSMZFOQFSRB-BJKOFHAPSA-N
Target: Ribosomal protein S6 kinase alpha-1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q15418
Assay: Phosphorylation Assay: HepG2 cells were obtained from ATCC and grown in DMEM supplemented with 10% fetal bovine serum. Cells were plated in 96-well plates at 35,000 cells/well and allowed to attach overnight at 37° C./5% CO2. Diluted compounds were then added at a final concentration of 0.5% DMSO. After 1.5 hour compound incubation, cells were stimulated with the addition of PMA (phorbol 12-myristate 13-acetate) at a final concentration of 100 ng/mL; the PMA stimulation was a 30-minute incubation at 37° C./5% CO2. After the 30-minute PMA stimulation, cells were washed with PBS and fixed in 3.7% formaldehyde in PBS at room temperature for 15-20 minutes. This was followed by another wash in PBS and then permeabilization in 100% MeOH at room temperature for 10-15 minutes. Following the permeabilization incubation, cells were washed in PBS/0.05% Tween-20, followed by a block in Odyssey blocking buffer (LI-COR Biosciences) for at least 1 hour. Antibodies to phosphorylated P90RSK(Ser380) (Cell Signaling #9335, rabbit monoclonal) and GAPDH (Fitzgerald 10R-G109a, mouse monoclonal) were added to the cells and incubated overnight at 4° C. pP90RSK(Ser380) antibody was used at a 1:250 dilution; GAPDH was used at a 1:10,000 dilution. After washing with PBS/0.05% Tween-20, the cells were incubated with fluorescently-labeled secondary antibodies (Anti-rabbit-Alexa Flour680, Invitrogen Cat#A21109; Anti-mouse-IRDye800CW, Rockland Inc. Cat#610-131-121) for 1 hour. Both secondary antibodies were used at a 1:1000 dilution. Cells were then washed and analyzed for fluorescence at both wavelengths using the Odyssey Infrared Imaging System (LI-COR Biosciences). Phosphorylated P90RSK(Ser380) signal was normalized to GAPDH signal.
Measurement: IC50 = 5.47 nM (pChEMBL 8.260000228881836)
Descriptors: MW 470.58, ALogP 3.86, HBA 7, HBD 2, PSA 84.31, QED 0.45
Compound (SMILES): FC1(F)Oc2ccc(CN3CCC(c4nnn5cnc6[nH]ccc6c45)CC3)cc2O1
InChIKey: IBELNCZALJQGIS-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 21 nM (pChEMBL 7.679999828338623)
Descriptors: MW 412.4, ALogP 3.31, HBA 7, HBD 1, PSA 80.57, QED 0.56
Compound (SMILES): COc1cccc(N2CC3C(c4ccccc4)(C2)NC(=N)N(C)S3(=O)=O)n1
InChIKey: ROOHNRUKVDMYSK-UHFFFAOYSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: Time-Resolved Endpoint Proteolysis Assay: Inhibitor IC50s at purified human autoBACE-2 were determined in a time-resolved endpoint proteolysis assay that measures hydrolysis of the QSY7-EISEVNLDAEFC-Eu-amide FRET peptide substrate (BACE-HTRF assay). BACE-mediated hydrolysis of this peptide results in an increase in relative fluorescence (RFU) at 620 nm after excitation with 320 nm light. Inhibitor compounds, prepared at 3x the desired final concentration in 1x BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO were pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1x BACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30 C. The assay was initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 uM for 4 uM for autoBACE-2) prepared in 1x BACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C.
Measurement: Ki = 26 nM (pChEMBL 7.579999923706055)
Descriptors: MW 387.47, ALogP 0.97, HBA 6, HBD 2, PSA 98.62, QED 0.81
Compound (SMILES): CCCS(=O)(=O)Nc1ccc(F)c(C(=O)c2c[nH]c3ncc(-c4ccc(F)nc4)cc23)c1F
InChIKey: UBJUBKFKARGYJS-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition of B-Raf V600E mutant (unknown origin) assessed as decrease in biotin-tagged MEK1 phosphorylation after 45 mins
Measurement: IC50 < 10 nM (pChEMBL 8)
Descriptors: MW 474.46, ALogP 4.42, HBA 5, HBD 2, PSA 104.81, QED 0.3
Compound (SMILES): Cc1cc(C#N)cnc1C(=O)Nc1cc(F)c(F)c([C@]2(CF)C[C@@H](C(F)(F)F)OC(N)=N2)c1
InChIKey: WWNBOKULGBAUQN-IFXJQAMLSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: In Vitro BACE Cell-Based Assay: The cell-based assay measures inhibition or reduction of Aβ40 in conditioned medium of test compound treated cells expressing amyloid precursor protein. Cells stably expressing Amyloid Precursor Protein (APP) were plated at a density of 40K cells/well in 96 well plates (Costar). The cells were cultivated for 24 hours at 37° C. and 5% CO2 in DMEM supplemented with 10% FBS. The test compounds were then added to cells in 10-point dose response concentrations with the starting concentration being either 100 uM or 10 uM. The compounds were diluted from stock solutions in DMSO and the final DMSO concentration of the test compounds on cells was 0.1%. After 24 h of incubation with the test compounds the supernatant conditioned media was collected and the Aβ 40 levels were determined using a sandwich ELISA.
Measurement: IC50 = 55 nM (pChEMBL 7.260000228881836)
Descriptors: MW 471.36, ALogP 3.62, HBA 6, HBD 2, PSA 113.39, QED 0.66
Compound (SMILES): CS(=O)(=O)c1ccc(C2CC(c3ccccc3)=NN2C(=O)Nc2ccc(F)cc2)cc1
InChIKey: KXOBYPUBQLDKPN-UHFFFAOYSA-N
Target: Voltage-dependent L-type calcium channel subunit alpha-1C (SINGLE PROTEIN), Homo sapiens
UniProt: Q13936
Assay: Inhibition of recombinant T-type calcium channel Cav1.2 (unknown origin) expressed in HEK293 cells assessed as inhibition of Kcl-induced calcium influx preincubated for 3 mins prior CaCl2 addition by Fluo-4-AM dye-based FLIPR assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 437.5, ALogP 4.61, HBA 4, HBD 1, PSA 78.84, QED 0.65
Compound (SMILES): O=C(CCCCCn1cc(-c2ccc3ncnc(Nc4ccc(OCc5cccc(F)c5)c(Cl)c4)c3c2)nn1)NO
InChIKey: JDBWHHHWEABNEC-UHFFFAOYSA-N
Target: Histone deacetylase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13547
Assay: Inhibition of HDAC1 (unknown origin) using acetylated peptide substrate preincubated for 15 mins followed by substrate addition measured after 60 mins by fluorescence analysis
Measurement: IC50 = 38.6 nM (pChEMBL 7.409999847412109)
Descriptors: MW 576.03, ALogP 6.07, HBA 9, HBD 3, PSA 127.08, QED 0.09
Compound (SMILES): O=C(CO)N(CCN1[C@@H]2CC[C@H]1C[C@@H](c1cccc(O)c1)C2)Cc1c(F)cccc1F
InChIKey: HIZQWWSJSYDBOP-REPLKXPHSA-N
Target: Delta-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41143
Assay: Displacement of [3H]-diprenorphine from human delta opioid receptor expressed in cell membranes
Measurement: Ki = 100 nM (pChEMBL 7)
Descriptors: MW 430.5, ALogP 3.4, HBA 4, HBD 2, PSA 64.01, QED 0.71
Compound (SMILES): COc1cc(OC)cc(N(CC#Cc2ccnc(N)n2)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: DBFMFHYDGXJVLJ-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 presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 2570.4 nM (pChEMBL 5.590000152587891)
Descriptors: MW 492.54, ALogP 3.61, HBA 10, HBD 1, PSA 117.1, QED 0.35
Compound (SMILES): CN(C)c1ccc2cccc(OCC(=O)NC[C@H](O)CN3CCc4ccccc4C3)c2n1
InChIKey: NQYRQXHXODTULO-NRFANRHFSA-N
Target: Protein arginine N-methyltransferase 5 (SINGLE PROTEIN), Homo sapiens
UniProt: O14744
Assay: Inhibition of PRMT5 in human Z138 cells assessed as decrease in di-methyl arginine levels after 96 hrs by Western blot method
Measurement: EC50 = 1001 nM (pChEMBL 6)
Descriptors: MW 434.54, ALogP 2.21, HBA 6, HBD 2, PSA 77.93, QED 0.57
Compound (SMILES): C[C@]12CC[C@H]3[C@@H](CCC4=CC(=O)CC[C@@]43C)C1=CC=C2c1cncnc1
InChIKey: VPLHRCLQGAXTGZ-FDTGPBLFSA-N
Target: Steroid 17-alpha-hydroxylase/17,20 lyase (SINGLE PROTEIN), Homo sapiens
UniProt: P05093
Assay: Inhibition of CYP17 in human testicular microsomes using 17alpha-hydroxypregnenolone as substrate assessed as decrease in DHEA levels after 90 mins by ELISA
Measurement: IC50 <= 11 nM (pChEMBL 7.959000110626221)
Descriptors: MW 346.47, ALogP 4.92, HBA 3, HBD 0, PSA 42.85, QED 0.72
Compound (SMILES): COc1cc(/C=N/NC(=O)[C@@H]2C[C@H]2c2cccc(Cl)c2)cc2cccnc12
InChIKey: RCQIXGLASLTZQE-BCEARJSMSA-N
Target: Transient receptor potential cation channel subfamily M member 5 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NZQ8
Assay: Inhibition of human TRPM5 transfected in HEK293 cells assessed as inhibition of KCl-stimulated membrane potential by FLIPR assay
Measurement: IC50 = 45 nM (pChEMBL 7.349999904632568)
Descriptors: MW 379.85, ALogP 4.15, HBA 4, HBD 1, PSA 63.58, QED 0.53
Compound (SMILES): CCCCN(CCCC)C(=O)c1nn(-c2ccc(C(=O)NS(=O)(=O)c3ccc4ccccc4c3)cc2C(=O)N2Cc3ccccc3C[C@H]2CN2CCN(CCO)CC2)c(C)c1Cl
InChIKey: YSUIAABABQYSSW-KDXMTYKHSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q07817
Assay: Fluorescence Resonance Energy Transfer Assay (FRET): The assays were performed in black flat-bottom 384-well plates. The final assay volume was 55 μl prepared from additions of Biotin-Bcl-xL (Bcl-xL: GENBANK® Accession No. Q07817), fluoresceinated 18-mer BIM peptide (NH2-YYANFEDGIRRLEQAIWI-[FAM]) (SEQ ID NO: 1), Streptavidin-Terbium FRET detection reagent, and test compounds in assay buffer consisting of 20 mM Sodium Phosphate, 1 mM EDTA, 50 mM NaCl, and 0.05% Pluronic Acid. The reaction was incubated at room temperature for 60 minutes. After 60 minutes, 5 μL of Streptavidin-Terbium FRET detection reagent (Perkin Elmer) was added to the reaction mixture and incubated at room temperature in the dark for 30 mins. The FRET signal generated by the reaction was detected on the Envision Plate Reader Inhibition data were calculated from FRET values generated by the no protein control reactions for 100% inhibition and vehicle-only reactions for 0% inhibition. The final concentration of reagents in the assay was 10 nM Bcl-xL, 5 nM fluo
Measurement: IC50 = 20 nM (pChEMBL 7.699999809265137)
Descriptors: MW 882.53, ALogP 6.33, HBA 10, HBD 2, PSA 148.39, QED 0.12
Compound (SMILES): CNC(=O)c1c(-c2ccc(C(F)F)cc2)oc2cc(N(C)S(C)(=O)=O)c(-c3ccc4c(n3)-c3cc5c(F)cccc5n3CO4)cc12
InChIKey: PFBQWEAGJPWFJS-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1b (isolate Japanese) (HCV)
UniProt: P26662
Assay: Inhbition Assay: To initiate an assay, replicon cells were plated in the presence of a dilution series of test compound in the absence of G418. Typically, the assays were performed in a 96-well plate formate for manual operation, or a 384-well plate format for automated assay. Replicon cells and compound were incubated for 96 hours. At the end of the assay, cells were washed free of media and compound, and the cells were then lysed. RNA was quantified indirectly through detection of replicon-encoded NS3/4A protein levels, through an ELISA-based assay with an antibody specific for NS3/4A.
Measurement: IC50 = 6.9 nM (pChEMBL 8.15999984741211)
Descriptors: MW 646.65, ALogP 6.97, HBA 7, HBD 1, PSA 106.67, QED 0.21
Compound (SMILES): Nc1ncnc2c1c(-c1ccc(Br)c(O)c1)nn2C1Cc2ccccc2C1
InChIKey: FZTMVHAUQSSXMI-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: Inhibition of PI3Kdelta (unknown origin) using biotinylated PIP2 as substrate in presence of streptavidin-APC by FRET assay
Measurement: IC50 = 8 nM (pChEMBL 8.100000381469727)
Descriptors: MW 422.29, ALogP 3.88, HBA 6, HBD 2, PSA 89.85, QED 0.51
Compound (SMILES): Cc1cccc(-c2ocnc2C(=O)Nc2cnn(CCOc3cnc4ccccc4n3)c2)c1
InChIKey: PHLDAHHMNAPBQI-UHFFFAOYSA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: Antagonist activity at human OX2R expressed in CHO cells assessed as orexin A-induced intracellular Ca2+ mobilization preincubated for 120 mins followed by orexin A addition by FLIPR assay
Measurement: IC50 = 1110 nM (pChEMBL 5.960000038146973)
Descriptors: MW 440.46, ALogP 4.12, HBA 8, HBD 1, PSA 107.96, QED 0.41
Compound (SMILES): NC(=O)c1nc(-c2ccncc2)sc1NC(=O)c1cn[nH]c1
InChIKey: HAVGDKYSPWKXRC-UHFFFAOYSA-N
Target: Inhibitor of nuclear factor kappa-B kinase subunit epsilon (SINGLE PROTEIN), Homo sapiens
UniProt: Q14164
Assay: IKKε Kinase Assay: The kinase assay is performed either as 384-well Flashplate assay (for e.g. Topcount measurement). 1 nM IKKε, 800 nM biotinylated IκBα(19-42) peptide (Biotin-C6-C6-GLKKERLLDDRHDSGLDSMKDEE) SEQ ID NO: 1 and 10 uM ATP (spiked with 0.3 uCi 33P-ATP/well) are incubated in a total volume of 50 ul (10 mM MOPS, 10 mM Mg-acetat, 0.1 mM EGTA, 1 mM Dithiothreitol, 0.02% Brij35, 0.1% BSA, 0.1% BioStab, pH 7.5) with or without test compound for 2 hours at 30° C. The reaction is stopped with 25 ul 200 mM EDTA. After 30 Min at room temperature the liquid is removed and each well washed thrice with 100 ul 0.9% sodium chloride solution. Non-specific reaction is determined in presence of 3 uM MSC2119074 (BX-795). Radioactivity is measured with Topcount (PerkinElmer).
Measurement: IC50 = 730 nM (pChEMBL 6.139999866485596)
Descriptors: MW 314.33, ALogP 1.28, HBA 6, HBD 3, PSA 126.65, QED 0.67
Compound (SMILES): COCC(=O)N[C@H]1CC[C@H](CCN2CCC(c3noc4cc(F)c(F)cc34)CC2)CC1
InChIKey: OCLIEDSOSAGNEM-JCNLHEQBSA-N
Target: D(3) dopamine receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35462
Assay: Radioligand Binding Assay: Aliquots of membrane preparations were thawed at RT, resuspended in assay buffer (D2, D3: 50 mM Tris-HCl, 120 mM NaCl, 5 mM MgCl2, 1 mM EDTA, 5 mM KCl, 1.5 mM CaCl2, pH=7.4; 5-HT2A: 50 mM Tris-HCl, 10 mM MgCl2, 1 mM EGTA, pH=7.4), homogenized with a Polytron for 20-30 sec at 12,000 rpm and adjusted to a final concentration of approximately 7.5 ug protein/well (D2, D3) and 15 ug protein/well (5-HT2A), respectively.The binding affinity (Ki) of the compounds was determined using radioligand binding. Membranes were incubated in a total volume of 200 ul with a fixed concentration of radioligand (final concentration approximately 0.7 nM [3H]-spiperone for D2, 0.5 nM [3H]-spiperone for D3, and 1.1 nM [3H]-ketanserin for 5-HT2A) and ten concentrations of test compound in ranging between 10 uM-0.1 nM for 1 h at RT. At the end of the incubation, the reaction mixtures were filtered on to unifilter 96-well white microplates with bonded GF/C filters.
Measurement: Ki = 32.67 nM (pChEMBL 7.489999771118164)
Descriptors: MW 435.52, ALogP 4, HBA 5, HBD 1, PSA 67.6, QED 0.71
Compound (SMILES): CC(C)N(C)CC[C@H](CSc1ccccc1)Nc1sc(S(=O)(=O)NC(=O)c2ccc(N3CCN(CC4=C(c5ccc(Cl)cc5)CCCC4)CC3)cc2)cc1[N+](=O)[O-]
InChIKey: KRIYPLQRPPASMS-PGUFJCEWSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q07817
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: Representative compounds were serially diluted in dimethyl sulfoxide (DMSO) starting at 50 μM (2× starting concentration; 10% DMSO) and 10 μL were transferred into a 384-well plate. Then 10 μL of a protein/probe/antibody mix was added to each well at final concentrations listed in TABLE 1. The samples are then mixed on a shaker for 1 minute and incubated for an additional 3 hours at room temperature. For each assay, the probe/antibody and protein/probe/antibody were included on each assay plate as negative and positive controls, respectively. Fluorescence was measured on the ENVISION plate reader (Perkin Elmer) using a 340/35 nm excitation filter and 520/525 (F-Bak peptide) and 495/510 nm (Tb-labeled anti-Histidine antibody) emission filters.
Measurement: Ki = 0.064 nM (pChEMBL 10.1899995803833)
Descriptors: MW 851.56, ALogP 8.88, HBA 11, HBD 2, PSA 128.13, QED 0.06
Compound (SMILES): CCC(CNC(=O)c1nc(C#N)c2cc(Oc3ccccc3)ccc2c1O)C(=O)O
InChIKey: GFXZDIYEOSVKGA-UHFFFAOYSA-N
Target: Egl nine homolog 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9GZT9
Assay: HIF-PH Assay: Ketoglutaric acid α-[1-14C]-sodium salt, alpha-ketoglutaric acid sodium salt, and HPLC purified peptide may be obtained from commercial sources, e.g., Perkin-Elmer (Wellesley Mass.), Sigma-Aldrich, and SynPep Corp. (Dublin Calif.), respectively. Peptides for use in the assay may be fragments of HIFα as described above or as disclosed in International Publication WO 2005/118836, incorporated by reference herein. HIF-pH, e.g., PHD1 (EGLN2) or PHD2 (EGLN1), can be expressed in, e.g., insect Hi5 cells, and partially purified, e.g., through a SP ion exchange chromatography column. Enzyme activity is determined by capturing 14CO2 using an assay described by Kivirikko and Myllyla (1982, Methods Enzymol. 82:245-304). Assay reactions contain 50 mM HEPES (pH 7.4), 100 μM α-ketoglutaric acid sodium salt, 0.30 mCi/mL ketoglutaric acid α-[1-14C]-sodium salt, 40 μM FeSO4, 1 mM ascorbate, 1541.8 units/mL Catalase, with or without 50 μM peptide substrate and various concentrations of compound of the invention. Reactions are initiated by addition of HIF-PH enzyme.
Measurement: IC50 = 5100 nM (pChEMBL 5.289999961853027)
Descriptors: MW 405.41, ALogP 3.44, HBA 6, HBD 3, PSA 132.54, QED 0.55
Compound (SMILES): NC(=O)c1cccc([C@H]2C[C@H]3CC[C@@H](C2)N3CCN(Cc2ccccc2)C(=O)[C@@H](O)CO)c1.O=C(O)C(F)(F)F
InChIKey: ZPLRMXXTPUHPSX-WTKDUOFSSA-N
Target: Delta-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41143
Assay: Displacement of [3H]-diprenorphine from human delta opioid receptor expressed in cell membranes
Measurement: Ki = 100 nM (pChEMBL 7)
Descriptors: MW 451.57, ALogP 1.88, HBA 5, HBD 3, PSA 107.1, QED 0.54
Compound (SMILES): CC(C)n1cc(-c2cc(-c3ccc(CN4CCOCC4)c(Cl)c3)cnc2N)nn1
InChIKey: NTYQUUSEPDYJKL-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: Kinase Activity Assay: The kinase reaction buffer consists of 50 mM Tris pH 8.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA, and 2 mM DTT. In the assay, 1 nM LRRK2 WT or 250 pM LRRK2 G2019S kinase is incubated with the test compound (typically at 0 to 30 μM) for 30 minutes before the kinase reaction is initiated by addition of 1.3 mM ATP and 0.4 μM fluorescein-LRRKtide. The reaction mixture (20 μl total volume) is incubated for 2 hours at 30 °C. before the reaction is terminated by addition of 10 mM EDTA and 1 nM terbium-labelled anti-phospho-LRRKtide antibody (final volume 20 μl). The mixture is further incubated for 30 minutes at RT. TR-FRET is measured by excitation of the terbium-donor with 340 nm light and subsequent (delay time 100 μs) measurement of terbium and fluorescein emission at 495 nm and 520 nm, respectively, over a time window of 1000 μs. The measurement is repeated 10 times for fluorescein and 10 times for terbium emission with a 2000 μs time window between repeats. TR-FRET measurements are performed on a Biomek Synergy plate. The TR-FRET signal is calculated as the emission-ratio at 520 nm over 495 nm.
Measurement: IC50 = 320 nM (pChEMBL 6.5)
Descriptors: MW 412.93, ALogP 3.66, HBA 7, HBD 1, PSA 82.09, QED 0.69
Compound (SMILES): COc1cc(O)cc(N(CCNC(C)C)c2ccc3ncc(-c4cn[nH]c4)nc3c2)c1.Cl
InChIKey: WDACLZKYOGTKNW-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 presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 418.5, ALogP 3.87, HBA 7, HBD 3, PSA 99.19, QED 0.4
Compound (SMILES): CC(C)c1nc2c(c(Nc3ccc(C(F)(F)F)cc3)n1)CCN(c1ncc([N+](=O)[O-])cc1C(F)(F)F)CC2
InChIKey: ISMANKQARRHHNX-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: O35433
Assay: Functional Assay: The assay was performed as described below, using HEK293 cells transfected with rat TRPV1 (rTRPV1/HEK293). These cells had a geneticin selection marker and were grown in Dulbecco's Modified Eagle Medium (DMEM, Gibco BRL) supplemented with 10% fetal bovine serum, penicillin/streptomycin (50 units/mL), and 500 μg/mL geneticin in 5% CO2 at 37° C. Cells were passaged frequently, every 3-5 days, to avoid overgrowth, depletion of essential medium components, or acidic medium exposure. Cells were passaged using a brief wash in 0.05% trypsin with 1 mM EDTA, followed by dissociation in divalent-free phosphate-buffered saline (Hyclone #SH30028.02). Dissociated cells were seeded onto poly-D-lysine coated black-walled 96-well plates (Biocoat; Becton Dickinson #354640) at about 40,000 cells per well and grown for approximately 1 day in culture medium to near confluency. The assay buffer was composed of 130 mM NaCl, 2 mM KCl, 2 mM MgCl2, 10 mM HEPES, 5 mM glucose, and either 2 mM or 20 μM CaCl2. On the day of the experiment, the culture medium was replaced with 2 mM calcium assay buffer using an automated plate washer (ELx405; Biotek, VT). The cells were incubated in 100 μL/well Fluo-3/AM (2 μM; TEFLabs #0116) with Pluronic F127 (100 μg/mL; Sigma #P2443) for 1 h at rt in the dark. After loading the cells, the dye solution was replaced with 50 μL/well of 20 μM calcium assay buffer using the ELx405 plate washer. Test compounds (50 μL/well) were added to the plate and incubated for 30 min. Intracellular Ca2+ levels were subsequently assayed using a Fluorometric Imaging Plate Reader (FLIPR™ instrument, Molecular Devices, CA) to simultaneously monitor Fluo-3 fluorescence in all wells (λexcitation=488 nm, λemission=540 nm) during challenge with agonist (capsaicin). The IC50 values were determined. Cells were challenged with 150 nM capsaicin and the fluorescence counts were captured following agonist addition at a sampling rate of 0.33 Hz. The contents of the wells were mixed 3 times (40 μL mix volume) immediately after the additions were made. Concentration dependence of block was determined by exposing each well of cells in duplicate rows of a 96 well plate to a serial dilution of test compound. The concentration series usually started at 10 Î¼M with a three-fold serial decrement in concentration. The magnitude of the capsaicin response was determined by measuring the change in fluo3 fluorescence before and 100 seconds after the addition of the agonist.
Measurement: IC50 = 170 nM (pChEMBL 6.769999980926514)
Descriptors: MW 540.47, ALogP 6.29, HBA 7, HBD 1, PSA 97.08, QED 0.23
Compound (SMILES): Cc1c(C(=O)Nc2ccc(F)c([C@]3(CF)C[C@@H](C(F)(F)F)OC(N)=N3)c2)nc2ccccn12
InChIKey: VBRNPACCSYLXRZ-MGPUTAFESA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: In Vitro BACE Cell-Based Assay: The cell-based assay measures inhibition or reduction of Aβ40 in conditioned medium of test compound treated cells expressing amyloid precursor protein. Cells stably expressing Amyloid Precursor Protein (APP) were plated at a density of 40K cells/well in 96 well plates (Costar). The cells were cultivated for 24 hours at 37° C. and 5% CO2 in DMEM supplemented with 10% FBS. The test compounds were then added to cells in 10-point dose response concentrations with the starting concentration being either 100 uM or 10 uM. The compounds were diluted from stock solutions in DMSO and the final DMSO concentration of the test compounds on cells was 0.1%. After 24 h of incubation with the test compounds the supernatant conditioned media was collected and the Aβ 40 levels were determined using a sandwich ELISA.
Measurement: IC50 = 888 nM (pChEMBL 6.050000190734863)
Descriptors: MW 467.4, ALogP 3.86, HBA 6, HBD 2, PSA 94.01, QED 0.57
Compound (SMILES): CC(C)N1CCN(C(=O)OC2CCN(c3ncc(CO)s3)CC2)CC1
InChIKey: ZFKNBJFIBYHAKV-UHFFFAOYSA-N
Target: Histamine H3 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5N1
Assay: Radioligand Binding Assay: The affinity of compounds of the invention to the H3 receptor can be assessed by measuring displacement of binding of the radioligand [3H]-N-Î±-Methylhistamine (PerkinElmer, #NET1027250UC) to membranes containing human H3 receptors (PerkinElmer, #ES-392-M400UA; GenBank: NM-007232.2; Hill S J et al, International Union of Pharmacology XIII. Classification of histamine receptors, Pharmacol Rev, 49(3), 253-278, 1997).Protocol: 1. Preparation of binding assay buffer (500 ml): 25 ml 1 M Tris-HCl pH 7.5 (Invitrogen, #15567-027), 2.5 ml 1 M MgCl2 (Sigma, #M1028-100ML), 472.5 ml ddH2O. 2. Compound serial dilution: Dilution was performed by BioTek Precision on compound dilution plate. Compound concentrations start at 5 or 10 uM, 10 point dose titrations with 3- or 5-fold serial dilutions. 3. Preparation of 2x membrane solution (25 ml): 1.25 ml human Histamine H3 receptor stock, 23.75 ml assay buffer. 4. Preparation of 2x solution of [3H]-N-Î±-methylhistamine (25 ml).
Measurement: Ki = 47 nM (pChEMBL 7.329999923706055)
Descriptors: MW 368.5, ALogP 1.77, HBA 7, HBD 1, PSA 69.14, QED 0.87
Compound (SMILES): O=C(O)CCCCCCC(=O)Nc1ccc(NCCOc2ccc3cc2COC/C=C/COCc2cccc(c2)-c2ccnc(n2)N3)cc1
InChIKey: XEDIJSKLNXIAPH-AATRIKPKSA-N
Target: Protein deacetylase HDAC6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UBN7
Assay: Inhibition of full length human recombinant N-terminal GST-tagged HDAC6 (1 to 1215 residues) expressed in sf9 cells preincubated with enzyme followed by fluorogenic Arg-His-Lys-Lys(Ac)-AMC substrate addition measured after 2 hrs by fluorescence assay
Measurement: IC50 = 2600 nM (pChEMBL 5.579999923706055)
Descriptors: MW 665.79, ALogP 7.34, HBA 9, HBD 4, PSA 143.93, QED 0.08
Compound (SMILES): Cc1cccc(-c2cc(N)nc3[nH]cc(C#N)c23)c1
InChIKey: KWTGXMXCSXFSJX-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: Inhibition of wild type recombinant human GST-tagged LRRK2 (970 to 2527 residues) expressed in baculovirus using fluorescein-LRRKtide as substrate after 2 hrs by TR-FRET assay
Measurement: IC50 = 21 nM (pChEMBL 7.679999828338623)
Descriptors: MW 248.29, ALogP 2.99, HBA 3, HBD 2, PSA 78.49, QED 0.69
Compound (SMILES): O=S(=O)(Nc1nccs1)c1ccc2c(c1)OCCN2c1ccccc1Cl
InChIKey: JIUSFZSCMYJTBW-UHFFFAOYSA-N
Target: Sodium channel protein type 5 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q14524
Assay: In Vitro Assay: HEK 293 cells stably transfected with either Nav 1.7 or with Nav 1.5 were recorded in population patch-clamp mode with the IonWorks® Quattro automated electrophysiology system in accordance with the manufacturer's specifications (Molecular Devices, LLC, Sunnyvale, Calif.). Sodium channel currents were measured in response to a train of depolarizations that induced successively greater inactivation. Cells were held at −110 mV for three seconds (Nav 1.7) or half a second (Nav 1.5) from a holding voltage of −15 mV, then put through a series of 26 pulses of 150 msec duration to −20 mV at a frequency of 5 Hz. Cells were then left unclamped for a period of 3 to 8 minutes while a single concentration of test compound was added. Cells were then reclamped and put through the same voltage protocol. Current at the end of the 26th pulse to −20 mV was subtracted from the peak current evoked by the 26th pulse to −20 mV to correct for leak current.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 407.9, ALogP 4.13, HBA 6, HBD 1, PSA 71.53, QED 0.7
Compound (SMILES): CCCS(=O)(=O)Nc1ccc(F)c(C(=O)c2c[nH]c3ncc(-c4ccc(O)nc4)cc23)c1F
InChIKey: MKVMMAUBKKDDSE-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition of B-Raf V600E mutant (unknown origin) assessed as decrease in biotin-tagged MEK1 phosphorylation after 45 mins
Measurement: IC50 < 10 nM (pChEMBL 8)
Descriptors: MW 472.47, ALogP 3.99, HBA 6, HBD 3, PSA 125.04, QED 0.35
Compound (SMILES): Cc1cc(Nc2nccc(Oc3ccc(NC(=O)Nc4cc(C(C)(C)C)nn4-c4ccc(=O)n(CCN5CCOCC5)c4)c4ccccc34)n2)cc2cn[nH]c12
InChIKey: XIFFATZVUNRCKL-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: c-Src and Syk Enzyme Inhibition Assay: The inhibitory activities of compounds of the invention against c-Src and Syk enzymes (Invitrogen), are evaluated in a similar fashion to that described hereinabove. The relevant enzyme (3000 ng/mL or 2000 ng/mL respectively, 2.5 uL) is incubated with the test compound (either 4 ug/mL, 0.4 ug/mL, 0.04 ug/mL, or 0.004 ug/mL, 2.5 uL each) for 2 hr at RT. The FRET peptides (8 uM, 2.5 uL), and appropriate ATP solutions (2.5 uL, 800 uM for c-Src, and 60 uM ATP for Syk) are then added to the enzymes/compound mixtures and incubated for 1 hr. Development reagent (protease, 5 uL) is added for 1 hr prior to detection in a fluorescence microplate reader (Varioskan Flash, ThermoFisher Scientific).
Measurement: IC50 = 3 nM (pChEMBL 8.520000457763672)
Descriptors: MW 753.87, ALogP 6.97, HBA 12, HBD 4, PSA 169.14, QED 0.11
Compound (SMILES): CN(C)CCOc1ccc(/C(=C(/CCCF)c2ccccc2)c2ccc(O)cc2)cc1.Cl
InChIKey: KXTIGEUSLGAAHZ-JTHROIFXSA-N
Target: Steroid hormone receptor ERR2 (SINGLE PROTEIN), Homo sapiens
UniProt: O95718
Assay: Inhibition of fluorescien-conjugated coactivator PGC1a binding to ERRbeta (unknown origin) after 1 hr by TR-FRET assay
Measurement: IC50 = 722 nM (pChEMBL 6.139999866485596)
Descriptors: MW 419.54, ALogP 6.04, HBA 3, HBD 1, PSA 32.7, QED 0.41
Compound (SMILES): CC1(C)CC2=NNC(=O)c3cc(F)cc4[nH]c(c2c34)C1=O
InChIKey: YADMFPWTDIQDHU-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P09874
Assay: Inhibition of PARP-1 in human HeLa cells incubated for 18 hrs in presence of H2O2
Measurement: EC50 = 7.6 nM (pChEMBL 8.119999885559082)
Descriptors: MW 285.28, ALogP 2.37, HBA 3, HBD 2, PSA 74.32, QED 0.78
Compound (SMILES): CN(C)CCCOc1ccc(-c2ccc3ncc4c(c3c2)n(C2(C)CCOCC2)c(=O)n4C)cn1
InChIKey: SIKMYPMDQGCEPL-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42336
Assay: Cellular Assay: Assay c) BT474 cells (human breast ductal carcinoma, ATCC HTB-20) were seeded into black 384 well plates (Costar, #3712) at a density of 5600 cells/well in DMEM containing 10% FBS and 1% glutamine and allowed to adhere overnight. The following morning compounds in 100% DMSO were added to assay plates by acoustic dispensing. After a 2 h incubation at 37° C. and 5% CO2, the medium was aspirated and the cells were lysed with a buffer containing 25 mM Tris, 3 mM EDTA, 3 mM EGTA, 50 mM sodium fluoride, 2 mM Sodium orthovanadate, 0.27 M sucrose, 10 mM β-glycerophosphate, 5 mM sodium pyrophosphate, 0.5% Triton X-100 and complete protease inhibitor cocktail tablets (Roche #04 693 116 001, used 1 tab per 50 ml lysis buffer). After 20 minutes, the cell lysates were transferred into ELISA plates (Greiner #781077) which had been pre-coated with an anti total-AKT antibody in PBS buffer and non-specific binding was blocked with 1% BSA in PBS containing 0.05% Tween 20. Plates were incubated over night at 4° C. The next day the plates were washed with PBS buffer containing 0.05% Tween 20 and further incubated with a mouse monoclonal anti-phospho AKT T308 for 2 h. Plates were washed again as above before addition of a horse anti-mouse-HRP conjugated secondary antibody. Following a 2 h incubation at r.t., plates were washed and QuantaBlu substrate working solution (Thermo Scientific #15169, prepared according to provider's instructions) was added to each well. The developed fluorescent product was stopped after 60 minutes by addition of Stop solution to the wells. Plates were read using a Tecan Satire plate reader using 325 nm excitation and 420 nm emission wavelengths respectively. Except where specified, reagents contained in the Path Scan Phospho AKT (Thr308) sandwich ELISA kit from Cell Signalling (#7144) were used in this ELISA assay.
Measurement: IC50 = 541 nM (pChEMBL 6.269999980926514)
Descriptors: MW 475.59, ALogP 3.81, HBA 8, HBD 0, PSA 74.41, QED 0.38