lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CCCCN(CCCC)C(=O)c1cc(C)n(-c2ccc(N[S+]([O-])c3ccc(NC(C)=O)c(Cl)c3)cc2C(=O)N2Cc3ccccc3C[C@H]2CO)n1
InChIKey: DANWOBJWEFZVQD-HDYGDMNUSA-N
Target: Apoptosis regulator Bcl-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P10415
Assay: Surface plasmon resonance (SPR): Methods for Determining IC50s.The present method includes utility of a Surface plasmon resonance (SPR)- based biosensor (Biacore GE Healthcare, Uppsala, Sweden) to characterize BCL-2 inhibitors.Biacore utilizes the phenomenon of surface plasmon resonance (SPR) to detect and measure binding interactions. In a typical Biacore experiment, one of the interacting molecules (ligand) is immobilized on a flexible dextran matrix while the interacting partner (analyte) is allowed to flow across that surface. A binding interaction results in an increase in mass on the sensor surface and a corresponding direct change in the refractive index of the medium in the vicinity of the sensor surface. Changes in refractive index or signal are recorded in resonance units (R.U.) Signal changes due to association and dissociation of complexes are monitored in a non-invasive manner, continuously and in real-time, the results of which are reported in the form of a sensorgram.
Measurement: IC50 = 254.5 nM (pChEMBL 6.590000152587891)
Descriptors: MW 733.33, ALogP 6.53, HBA 8, HBD 3, PSA 142.86, QED 0.13
Compound (SMILES): CC1(C)CCC(CN2CCN(c3ccc(C(=O)NS(=O)(=O)c4cnc(OCC5CCC(C)(O)CC5)c(Cl)c4)c(Oc4cnc5[nH]ccc5c4)c3)CC2)=C(c2ccc(Cl)cc2)C1
InChIKey: JHPHSPYSBGEZFM-UHFFFAOYSA-N
Target: Apoptosis regulator Bcl-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P10415
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: Representative compounds were serially diluted in dimethyl sulfoxide (DMSO) starting at 50 uM (2x starting concentration; 10% DMSO) and 10 uL were transferred into a 384-well plate. Then 10 uL of a protein/probe/antibody mix was added to each well at final concentrations listed in TABLE 1. Protein:GST-Bcl-2, Probe: F-Bak Peptide Probe Acetyl-GQVGRQLAIIGDK(6-FAM)INR-amide(SEQ ID NO: 1), Protein(nM): 1, Probe (nM): 100, Antibody: Tb-anit-GST, Antibody (nm): 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 = 0.034 nM (pChEMBL 10.470000267028809)
Descriptors: MW 887.93, ALogP 9.28, HBA 10, HBD 3, PSA 149.98, QED 0.11
Compound (SMILES): C[C@@H](NC(=O)c1nn(-c2ccc(Cl)cc2)c2c1CN(C(=O)c1nc[nH]n1)C/C2=C\c1ccc(F)cc1)c1ccccc1
InChIKey: QSOFEUWMAZQPQA-QTCOBDOFSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: Inverse agonist activity at human CB1 receptor transfected in HEK293 cells assessed as increase of forskolin-induced cAMP production incubated for 30 mins followed by d2 labeled cAMP addition measured after 60 mins by HTRF assay
Measurement: EC50 = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 582.04, ALogP 5.47, HBA 6, HBD 2, PSA 108.8, QED 0.28
Compound (SMILES): CC(C)c1ccc(-c2nnc(-c3cc(Cl)c([C@]4(C)CS(=O)(=O)N(C)C(=N)N4)s3)o2)cc1
InChIKey: KLTFUOZRHPUBQI-FQEVSTJZSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: Time-Resolved Endpoint Proteolysis Assay: Inhibitor compounds, prepared at 3× the desired final concentration in 1×BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO are pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1×BACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30° C. The assay is initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 μM for 4 μM for autoBACE-2) prepared in 1×BACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C. DMSO is present at 5% final concentration in the assay. Following laser excitation of sample wells at 320 nm, the fluorescence signal at 620 nm is collected for 400 ms following a 50 μs delay on a RUBYstar HTRF plate reader (BMG Labtechnologies).
Measurement: Ki = 889.4 nM (pChEMBL 6.050000190734863)
Descriptors: MW 480.02, ALogP 4.26, HBA 7, HBD 2, PSA 112.18, QED 0.58
Compound (SMILES): Cc1cc2ccc1[C@@H](C)COC(=O)Nc1cc(F)c(O[C@@H](C)CO)c(c1)CN(C)C(=O)[C@@H]2Nc1ccc2c(N)nccc2c1
InChIKey: NINDJBINKRLZFB-QWQFASRJSA-N
Target: Coagulation factor VII (SINGLE PROTEIN), Homo sapiens
UniProt: P08709
Assay: In Vitro Assay: Tissue kallikrein-1 (37° C) activity was determined in reactions containing 0.05 nM enzyme and 90 μM substrate (H-D-Val-Leu-Arg-AFC) in buffer (0.1M sodium phosphate pH 7.4, 0.2 M NaCl, 0.5% PEG 8000, and 1% DMSO). Assays were performed using 96-well microtiter plates (COSTAR 3600, CORNING, NY, USA) and a thermostatic temperature controlled plate reader (SPECTRAMAX® Gemini, Molecular Devices, Sunnyvale, Calif., USA). Fluorescence was monitored using 400 nm excitation and 505 nm emission wavelengths.
Measurement: Ki = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 601.68, ALogP 5.5, HBA 8, HBD 4, PSA 139.04, QED 0.24
Compound (SMILES): CCN(CC)c1nc(NCc2ccc(NC(=O)C3CCN(Cc4ccc(F)cc4)CC3)cc2)c2cc(C(F)(F)F)ccc2n1
InChIKey: FMQIPZPIEGHYNH-UHFFFAOYSA-N
Target: TCF4/beta-catenin (PROTEIN COMPLEX), Homo sapiens
UniProt: P35222
Assay: Inhibition of TCF-4/beta catenin in human Tcf22C11 cells expressing Tcf-4/SV40 binding sites driving firefly luciferase reporter gene after 24 hrs by luminometer
Measurement: IC50 > 4107 nM (pChEMBL 5.386000156402588)
Descriptors: MW 608.68, ALogP 7.1, HBA 6, HBD 2, PSA 73.39, QED 0.19
Compound (SMILES): CN(C)S(=O)(=O)c1ccc(OC/C(=C/F)CN)cc1
InChIKey: QESDALNWZLIPAA-JXMROGBWSA-N
Target: Amine oxidase [copper-containing] 3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q16853
Assay: Recombinant SSAO/VAP-1 Inhibtion Assay: Briefly, HMEC cell expressing human SSAO/VAP-1 were grown in several 10 cm petri dishes, once the cells reached 100% confluency, cells were harvested and homogenates were prepared. Cells were washed twice with 5 mL of chilled HES buffer (20 mM HEPES, 1 mM EDTA, 250 mM sucrose, pH 7.4). HES buffer containing 1× protease inhibitor (Sigma Aldrich) and added and cells were incubated on ice for 3 min. Buffer was removed and cells were scraped and transferred to a centrifuge tube. Cell lysates were prepared by passing through 23 G needle for, 10 times and followed by 27 G needle for 10 times. Alternatively the cell lysates were prepared by using IKA Ultra-Turrax T 10 homogenizer for 3 min for every 10 mL of cell suspensions. Cells were then spun for 5 min at 300×g. The clear supernatant was transferred to new centrifuge tube and stored at −80 °C. until colorimetric assay was performed. Prior to the assay, 0.5 mM pargyline was added in order to inhibit any residue MAO activities. Briefly, 50 μL of cell lysate was incubated with test compounds for 30 min at 37 °C. Reaction mixtures were added and kinetic was read as described in detail in Example 5: Briefly, in a standard 96 well plate assay 50 μL of purified human SSAO/VAP-1 (0.25 μg/mL) in 0.1 M NaPO4 buffer (pH 7.4) was added into each well. Test compounds were dissolved in DMSO and tested in a Concentration Response Curve (CRC) with 4-9 data points, typically in the micromolar or nanomolar range after incubation with human SSAO/VAP-1 for 30 min at 37 °C. After 30 min incubation, 50 μL of the reaction mixture containing 600 μM benzylamine (Sigma Aldrich), 120 μM Amplex Red (Sigma Aldrich) and 1.5 U/mL horseradish peroxidase (Sigma Aldrich) prepared in 0.1 M NaPO4 buffer (pH 7.4) were added to the corresponding well. The fluorescence unit (RFU) was read every 2.5 min for 30 min at 37 °C. excitation 565 nm and emission 590 (Optima; BMG labtech). The slope of the kinetics for each well was calculated using MARS data analysis software (BMG labtech) and this value was used to deduce the IC50 value (Dotmatics).
Measurement: IC50 < 100000 nM (pChEMBL 4)
Descriptors: MW 288.34, ALogP 1.13, HBA 4, HBD 1, PSA 72.63, QED 0.85
Compound (SMILES): Cn1cc(-c2cc(-c3ccc(F)nc3)ccc2OCC2CC2)c2cc[nH]c2c1=O
InChIKey: OMJNRHJWPMTRLF-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 = 422 nM (pChEMBL 6.380000114440918)
Descriptors: MW 389.43, ALogP 4.52, HBA 4, HBD 1, PSA 59.91, QED 0.51
Compound (SMILES): NS(=O)(=O)c1ccc(SCc2ccc3ccccc3c2)s1
InChIKey: FBVRPMPAPFOBSJ-UHFFFAOYSA-N
Target: Carbonic anhydrase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: Q16790
Assay: Inhibition of human recombinant transmembrane tumor associated carbonic anhydrase 9 preincubated for 10 mins prior to testing by stopped-flow CO2 hydration assay
Measurement: Ki = 8.3 nM (pChEMBL 8.079999923706055)
Descriptors: MW 335.48, ALogP 3.84, HBA 4, HBD 1, PSA 60.16, QED 0.74
Compound (SMILES): Cc1ccccc1-n1c(Cn2nc(C(N)=O)c3c(N)ncnc32)cc2cccc(C)c2c1=O
InChIKey: ZBMSILUYNLJEMN-UHFFFAOYSA-N
Target: PI3-kinase p110-delta/p85-alpha (PROTEIN COMPLEX), Homo sapiens
UniProt: O00329
Assay: Inhibition of N-terminal His6-tagged recombinant full-length human p110delta/untagged recombinant full length p85alpha expressed in baculovirus infected Sf21 cells using phosphatidylinositol as substrate in presence of gamma-32P-ATP after 5 mins by thin layer chromatographic method
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 439.48, ALogP 2.48, HBA 8, HBD 2, PSA 134.71, QED 0.44
Compound (SMILES): Cc1ccc(C(=O)N2CCC(N3CC(CC#N)(n4cc(-c5ncnc6[nH]ccc56)cn4)C3)CC2)s1
InChIKey: SIPLUFHWHHQVCL-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Inhibition of human N-terminal His-tagged JAK2 catalytic domain (828 to 1132 residues) expressed in baculovirus infected insect cells assessed as decrease in phosphorylation of biotinylated peptide substrate after 1 hr in presence of 1 mM ATP by HTRF method
Measurement: IC50 <= 10 nM (pChEMBL 8)
Descriptors: MW 486.61, ALogP 3.42, HBA 8, HBD 1, PSA 106.73, QED 0.46
Compound (SMILES): CNC(=N)NCCC[C@H](NC(=O)[C@H](CC(C)C)NC(=O)NNC(=O)[C@H](Cc1cccc(F)c1)NC(=O)[C@@H](NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CCC(=O)O)NC(=O)[C@@H](Cc1ccc(O)cc1)NC(C)=O)[C@@H](C)O)C(=O)N[C@@H](Cc1c[nH]c2ccccc12)C(N)=O
InChIKey: BNSPMRCFHAFQER-WBDBVSIISA-N
Target: KiSS-1 receptor (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q924U1
Assay: Agonist activity at rat KISS1R expressed in CHO/dhfr cells assessed as increase in intracellular Ca2+ levels measured for 180 secs by Fluo 3-AM dye based FLIPR assay
Measurement: EC50 = 20 nM (pChEMBL 7.699999809265137)
Descriptors: MW 1259.36
Compound (SMILES): CN(C)C(=O)CCCN(C)C1CCN(C(=O)Cn2cc(NC(=O)c3cnn4cccnc34)c(-c3cc(Cl)ccc3OC(F)F)n2)CC1
InChIKey: XVNALBBKCPINNQ-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: JAK Enzyme Assay: The activity of the isolated recombinant JAK1 and JAK2 kinase domain was measured by monitoring phosphorylation of a peptide derived from JAK3 (Val-Ala-Leu-Val-Asp-Gly-Tyr-Phe-Arg-Leu-Thr-Thr, fluorescently labeled on the N-terminus with 5-carboxyfluorescein) using the Caliper LabChip® technology (Caliper Life Sciences, Hopkinton, Mass.). To determine inhibition constants (Ki), compounds were diluted serially in DMSO and added to 50 μL kinase reactions containing purified enzyme (1.5 nM JAK1, or 0.2 nM JAK2), 100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 1.5 μM peptide substrate, ATP (25 μM), 10 mM MgCl2, 4 mM DTT at a final DMSO concentration of 2%. Reactions were incubated at 22° C. in 384-well polypropylene microtiter plates for 30 minutes and then stopped by addition of 25 μL of an EDTA containing solution (100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 150 mM EDTA), resulting in a final EDTA concentration of 50 mM. After termination of the kinase reaction, the proportion of phosphorylated product was determined as a fraction of total peptide substrate using the Caliper LabChip® 3000 according to the manufacturer's specifications.
Measurement: Ki = 1.91 nM (pChEMBL 8.720000267028809)
Descriptors: MW 672.14, ALogP 3.89, HBA 10, HBD 1, PSA 130.2, QED 0.24
Compound (SMILES): CCN1CCN(c2noc(COc3cc4ncnc(Nc5ccc(Cl)c(Cl)c5)c4cc3OC)n2)CC1
InChIKey: FGHXOSBHGQKMBX-UHFFFAOYSA-N
Target: Ephrin type-A receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29317
Assay: Inhibition of recombinant human N-terminal His6-tagged EphA2 (N598 to R890 residues) expressed in baculovirus using poly-EY as substrate after 180 mins by luciferase coupled chemiluminescence assay
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 530.42, ALogP 4.79, HBA 10, HBD 1, PSA 101.67, QED 0.34
Compound (SMILES): O=C(Nc1ccc(C2CCCCN2)cc1)c1ccc(Cl)cc1
InChIKey: ZEDVYTYZHLXBDQ-UHFFFAOYSA-N
Target: Trace amine-associated receptor 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q923Y9
Assay: Radioligand Binding Assay: HEK-293 cells stably expressing rat TAAR1 were maintained at 37° C. and 5% CO2 in DMEM high glucose medium, containing fetal calf serum (10%, heat inactivated for 30 min at 56° C.), penicillin/streptomycin (1%), and 375 ug/ml geneticin (Gibco). Cells were released from culture flasks using trypsin/EDTA, harvested, washed twice with ice-cold PBS (without Ca2+ and Mg2+), pelleted at 1,000 rpm for 5 min at 4° C., frozen and stored at -80° C. Frozen pellets were suspended in 20 ml HEPES-NaOH (20 mM, pH 7.4) containing 10 mM EDTA and homogenized with a Polytron (PT 6000, Kinematica) at 14,000 rpm for 20 s. The homogenate was centrifuged at 48,000Ã¿g for 30 min at 4° C. Subsequently, the supernatant was removed and discarded, and the pellet resuspended in 20 ml HEPES-NaOH (20 mM, pH 7.4) containing 0.1 mM EDTA using the Polytron (20 s at 14,000 rpm). This procedure was repeated and the final pellet resuspended in HEPES-NaOH containing 0.1 mM EDTA and homogenized using the Polytron. Typically, aliquots of 2 ml membrane portions were stored at -80° C. With each new membrane batch the dissociation constant (Kd) was determined via a saturation curve. The TAAR1 radioligand 3[H]â¿¿(S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine (described in WO 2008/098857) was used at a concentration equal to the calculated Kd value, that was usually around 2.3 nM, resulting in the binding of approximately 0.2% of the radioligand and a specific binding representing approximately 85% of the total binding. Nonspecific binding was defined as the amount of 3[H]â¿¿(S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine bound in the presence of 10 uM unlabeled ligand. All compounds were tested at a broad range of concentrations (10 pM to 10 uM) in duplicates. The test compounds (20 ul/well) were transferred into a 96 deep well plate (TreffLab), and 180 ul of HEPES-NaOH (20 mM, pH 7.4) containing MgCl2 (10 mM) and CaCl2 (2 mM) (binding buffer), 300 ul of the radioligand 3[H]â¿¿(S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine at a concentration of 3.3xKd in nM and 500 ul of the membranes (resuspended at 50 ug protein per ml) added. The 96 deep well plates were incubated for 1 hr at 4° C. Incubations were terminated by rapid filtration through Unifilter-96 plates (Packard Instrument Company) and glass filters GF/C (Perkin Elmer) presoaked for 1 hr in polyethylenimine (0.3%) and washed 3 times with 1 ml of cold binding buffer. After addition of 45 ul of Microscint 40 (PerkinElmer) the Unifilter-96 plate was sealed and after 1 hr the ratioactivity counted using a TopCount Microplate Scintillation Counter (Packard Instrument Company).
Measurement: Ki = 629.8 nM (pChEMBL 6.199999809265137)
Descriptors: MW 314.82, ALogP 4.41, HBA 2, HBD 2, PSA 41.13, QED 0.88
Compound (SMILES): CCC(=O)Nc1cc2c(Nc3ccc4c(ccn4Cc4ccccc4)c3)ncnc2cc1OC
InChIKey: GAYUJMWHGBAPKD-UHFFFAOYSA-N
Target: Receptor tyrosine-protein kinase erbB-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P04626
Assay: Irreversible inhibition of GST-tagged ERBB2 (unknown origin) (Ile-675 to Val-1256 residues) expressed in baculovirus infected Sf9 insect cells assessed as reduction in Glu/Tyr copolymer phosphorylation after 6 mins by ELISA
Measurement: IC50 = 124 nM (pChEMBL 6.909999847412109)
Descriptors: MW 451.53, ALogP 5.73, HBA 6, HBD 2, PSA 81.07, QED 0.33
Compound (SMILES): COc1cc(F)cc(-c2nn(C3Cc4ccccc4C3)c3ncnc(N)c23)c1
InChIKey: GQNBIEAGAUZGAD-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 = 2790 nM (pChEMBL 5.550000190734863)
Descriptors: MW 375.41, ALogP 3.56, HBA 6, HBD 1, PSA 78.85, QED 0.59
Compound (SMILES): CCN(CC)CCCC(=O)Nc1nc2cc(C3(O)c4ccccc4C(=O)N3Cc3ccccc3)ccc2[nH]1
InChIKey: YRZZQUOMZIVMFU-UHFFFAOYSA-N
Target: RAF proto-oncogene serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P04049
Assay: Inhibition of c-Raf1 (unknown origin) pre-incubated for 30 mins by luciferase-luciferin-coupled chemiluminescence assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 511.63, ALogP 4.47, HBA 5, HBD 3, PSA 101.56, QED 0.29
Compound (SMILES): CCOc1ncc2ccccc2c1C(=O)N1[C@@H]2CC[C@H]1[C@H](COc1ccccn1)C2
InChIKey: DQXRZOLGSQYVPN-NSHGMRRFSA-N
Target: Orexin/Hypocretin receptor type 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O43613
Assay: Radioligand Binding Assay: Human Embryonic Kidney 293 cells (HEK293) stably expressing rat orexin 1 receptor (Genebank accession number NM_001525) or Chinese ovary cells (CHO) stably expressing human orexin 1 receptor (Genebank accession number NM_001526) were grown to confluency in DMEM (Hyclone, cat #SH30022), 10% FBS, 1x Pen/Strep, 1x sodium pyruvate, 10 mM HEPES, 600 ug/mL G418 and DMEM/F12 (Gibco, Cat #11039), 10% FBS, 1x Pen/Strep, 600 ug/mL G418 media, respectively on 150 cm^2 tissue culture plates, washed with 5 mM EDTA in PBS (HyClone Dulbecco's Phosphate Buffered Saline 1x with Calcium and Magnesium, Cat #SH30264.01, hereafter referred to simply as PBS) and scraped into 50 ml tubes. After centrifugation (2KxG, 5 min at 4° C.), the supernatant was aspirated and the pellets frozen and stored at -80\°C. Cells were resuspended in PBS in the presence of 1 tablet of protease inhibitor cocktail (Roche, Cat. #11836145001) per 50 mL. Each cell pellet from a 15 cm plate was resuspended in 10 mL, stored on ice, and homogenized for 45 sec prior to addition to the reactions. Competition binding experiments in 96 well polypropylene plates were performed using [3H]-SB674042 (Moraveck Corporation, specific activity=35.3 Ci/mmol), diluted to a 10 nM concentration in PBS (4 nM final). Compounds were solubilized in 100% DMSO (Acros Organics, Cat. #61042-1000) and tested over a range of 7 concentrations (from 0.1 nM to 10 uM). The final concentration of DMSO in the reactions is equal to or less than 0.1%. Total and nonspecific binding was determined in the absence and presence of 10 uM almorexant. The total volume of each reaction is 200 uL (20 uL of diluted compounds, 80 uL of [3H]-(1-(5-(2-fluoro-phenyl)-2-methyl-thiazol-4-yl)-1-((S)-2-(5-phenyl-(1,3,4)oxadiazol-2-ylmethyl)-pyrrolidin-1-yl)-methanone) diluted in PBS and 100 uL of the cell suspension). Reactions were run for 60 min at room temperature and terminated by filtration through GF/C filter plates (PerkinElmer, Cat. #6005174) presoaked in 0.3% polyethylenimine using the cell harvester (PerkinElmer Filtermate). The plates were washed 3 times by aspirating 30 ml PBS through the plates. Plates were dried in 55° C. oven for 60 min, scintillation fluid was added, and the radioactivity was counted on a Topcount (Packard).
Measurement: Ki = 7 nM (pChEMBL 8.149999618530273)
Descriptors: MW 403.48, ALogP 4.1, HBA 5, HBD 0, PSA 64.55, QED 0.62
Compound (SMILES): O=c1cc(NC2CCN(S(=O)(=O)C(F)(F)F)CC2)c2cc(C(c3ccc(Cl)cc3)c3ccc(Cl)cc3)cc(NC3CCN(S(=O)(=O)C(F)(F)F)CC3)c2[nH]1
InChIKey: RKRCBMVEJROYQW-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 840.7, ALogP 7.47, HBA 7, HBD 3, PSA 131.68, QED 0.12
Compound (SMILES): Cc1nsc(-c2nnc3n2CCN(C(=O)c2ccc(F)cc2)C3C)n1
InChIKey: PPSNFPASKFYPMN-UHFFFAOYSA-N
Target: Neuromedin-K receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P29371
Assay: Competitive Binding Assay: The affinity of compounds of the invention for the human NK-3 receptor was determined by measuring the ability of compounds of the invention to competitively and reversibly displace a well-characterized NK-3 radioligand in a concentration-dependent manner.3H-SB222200 Binding Competition Assay with Human NK-3 ReceptorThe ability of compounds of the invention to inhibit the binding of the NK-3 receptor selective antagonist 3H-SB222200 was assessed by an in vitro radioligand binding assay. Membranes were prepared from Chinese hamster ovary recombinant cells stably expressing the human NK-3 receptor. The membranes were incubated with 5 nM 3H-SB222200 (ARC) in a HEPES 25 mM/NaCl 0.1M/CaCl2 1 mM/MgCl2 5 mM/BSA 0.5%/Saponin 10 μg/ml buffer at pH 7.4 and various concentrations of compounds of the invention. The amount of 3H-SB222200 bound to the receptor was determined after filtration by the quantification of membrane associated radioactivity using the TopCount-NXT reader (Packard).
Measurement: Ki = 42 nM (pChEMBL 7.380000114440918)
Descriptors: MW 358.4, ALogP 2.46, HBA 7, HBD 0, PSA 76.8, QED 0.7
Compound (SMILES): CN1CC(Cn2cc(-c3ccc(F)c(C(F)(F)F)c3)nc2C2CCN(c3ncnc(N)c3C(N)=O)CC2)C1
InChIKey: GFWUCLLYVKRKSO-UHFFFAOYSA-N
Target: RAC-alpha serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P31749
Assay: AKT Enzyme Assay: A TTP Mosquito liquid handling instrument is used to place 125 nl of the appropriate concentration of inhibitor in 100% DMSO (for a dose response curve calculation) into each well of a 384-well plate. To this reaction components are added to a final volume of 12.5 .mu.l: 0.1 ng/.mu.l His-AKT (Full Length), (Invitrogen, Part # P2999, Lot #641228C). 160 uM ATP (Fluka, 02055) 1 mM DTT (Sigma, D0632) 1 mM MgCl2 (Sigma, M1028) 1 .mu.M substrate peptide (sequence FITC-AHA-GRPRTSSFAEG-NH2), synthesized by Tufts Peptide Synthesis service. 100 mM HEPES pH 7.5 (Calbiochem, 391338)0.015% Brij-35 (Sigma, B4184)The reaction is incubated for 90 min at 25 C, and then stopped by the addition of 70 .mu.l of Stop buffer (100 mM HEPES pH 7.5, 0.015% Brij-35, 10 mM EDTA (Sigma, E7889)).The plate is read on a Caliper LC 3000 in an Off-Chip mobility shift assay format, using the following parameters for a 12-sipper chip: screening pressure -2.3 psi, upstream voltage -500, and downstream voltage -3000.
Measurement: IC50 = 81 nM (pChEMBL 7.090000152587891)
Descriptors: MW 532.55, ALogP 3.12, HBA 8, HBD 2, PSA 119.19, QED 0.47
Compound (SMILES): CC[C@@]1(c2ccccc2)NC(=O)N(Cc2ccccc2)C1=O
InChIKey: ZMZDHUHMXXALFX-SFHVURJKSA-N
Target: Cytochrome P450 2C19 (SINGLE PROTEIN), Homo sapiens
UniProt: P33261
Assay: Inhibition of CYP2C19 in human liver microsomes using S-mephenytoin as substrate preincubated for 10 mins followed by NADPH addition measured after 10 mins by LC/MS/MS analysis
Measurement: IC50 = 211 nM (pChEMBL 6.679999828338623)
Descriptors: MW 294.35, ALogP 3.04, HBA 2, HBD 1, PSA 49.41, QED 0.88
Compound (SMILES): O=C(NO)c1ccc(CNC(=O)c2nc(-c3nccs3)sc2C2CC2)cc1
InChIKey: HPAOMQPZUJIWDB-UHFFFAOYSA-N
Target: Histone deacetylase 3 (SINGLE PROTEIN), Homo sapiens
UniProt: O15379
Assay: Enzyme Assay: Enzyme activity of HDAC1,3 is determined using the substrate Ac-Lys-Tyr-Lys(Ac)-AMC while the enzyme activity of HDAC6 is assayed using the substrate Boc-lys(Ac)-AMC. The reaction is carried out in flat-bottom, 96-well, or 384-well microplates. After the substrate was deacetylated by HDACs, the product AMC which was obtained from hydrolysis by trypsin can generate fluorescence signal. Measurement is taken in using an multilabe plate reader a 355 nm excitation filter and a 460 nm emission filter. The initial rate of fluorescence should accurately reflect the rate of product formation and enzyme activity. Sample processing: The sample was dissolved in DMSO and kept at a low temperature. DMSO in the final system is limited under a low concentration which won't affect the enzyme activity.
Measurement: IC50 = 30 nM (pChEMBL 7.519999980926514)
Descriptors: MW 400.49, ALogP 3.19, HBA 7, HBD 3, PSA 104.21, QED 0.44
Compound (SMILES): Cc1ccc(-c2ccc(N3CCc4cccc(C(=O)Nc5nc6ccccc6s5)c4C3)nc2C(=O)O)cc1Oc1ccccc1
InChIKey: PALSTDWPMINOGA-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: FL5.12 Cellular Assay: The efficacy of the compounds of Formula (I) can also be determined in cell-based killing assays using a variety of cell lines and mouse tumor models. For example, their activity on cell viability can be assessed on a panel of cultured tumorigenic and non-tumorigenic cell lines, as well as primary mouse or human cell populations. In one exemplary set of conditions, mouse FL5.12 cells transfected with Bcl-XL were cultured under standard conditions in RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3). For assaying the compound activity, the cells were exchanged into an IL-3-depleted deprivation media, which was identical to the growth media except for the absence of FBS and WEHI-3B conditional media, for 2 days. Then the cells were exchanged to 3% FBS assay media (RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3).
Measurement: EC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 612.71, ALogP 7.97, HBA 7, HBD 2, PSA 104.65, QED 0.19
Compound (SMILES): CC(C)CC1(Cn2cc(-c3ccc(N4CCc5cccc(C(=O)Nc6nc7ccccc7s6)c5C4)nc3C(=O)O)cn2)CCCCC1
InChIKey: MVQZRHWCVRNVCZ-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: FL5.12 Cellular Assay: The efficacy of the compounds of Formula (I) can also be determined in cell-based killing assays using a variety of cell lines and mouse tumor models. For example, their activity on cell viability can be assessed on a panel of cultured tumorigenic and non-tumorigenic cell lines, as well as primary mouse or human cell populations. In one exemplary set of conditions, mouse FL5.12 cells transfected with Bcl-XL were cultured under standard conditions in RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3). For assaying the compound activity, the cells were exchanged into an IL-3-depleted deprivation media, which was identical to the growth media except for the absence of FBS and WEHI-3B conditional media, for 2 days. Then the cells were exchanged to 3% FBS assay media (RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3).
Measurement: EC50 = 124 nM (pChEMBL 6.909999847412109)
Descriptors: MW 648.83, ALogP 8.06, HBA 8, HBD 2, PSA 113.24, QED 0.17
Compound (SMILES): Cn1ccc2cc(OCCCN3CCN(c4cccc5sccc45)CC3)ccc2c1=O
InChIKey: FQLPASRHPPCIMN-UHFFFAOYSA-N
Target: 5-hydroxytryptamine receptor 2A (SINGLE PROTEIN), Rattus norvegicus
UniProt: P14842
Assay: Serotonin 5-HT2A Receptor Binding Assay: The assay was performed according to the method by Leysen J E et al. (Leysen J E, Niemegeers C J E, Van Nueten J M and Laduron P M. [3H] Ketanserin (R 41 468), a selective 3H-ligand for serotonin 2 receptor binding sites. Mol. Pharmacol., 1982, 21: 301-314). The binding assay was performed using 40 ul of the membrane specimen, 20 ul of [3H]-Ketanserin (final concentration 1 to 3 nM), 20 ul of a test drug and 50 mM Tris-hydrochloric acid buffer (pH 7.4) so that the total amount was 200 ul (final dimethylsulfoxide concentration 1%). The reaction was performed at 37° C. for 20 minutes and terminated by conducting suction filtration with a cell harvester on a glass fiber filter plate.The filter plate made of glass fiber was washed with 50 mM Tris-hydrochloric acid buffer (pH 7.4), and after dried, a microplate liquid scintillation cocktail was added and the radioactivity was measured with a microplate scintillation counter.
Measurement: Ki = 3.2 nM (pChEMBL 8.489999771118164)
Descriptors: MW 433.58, ALogP 4.34, HBA 6, HBD 0, PSA 37.71, QED 0.42
Compound (SMILES): COc1ccc(Cl)c(N2CCC3(CCC(CC(=O)O)CC3)CC2)c1
InChIKey: YIXZOCHHRTUZKL-UHFFFAOYSA-N
Target: Free fatty acid receptor 4 (SINGLE PROTEIN), Mus musculus
UniProt: Q7TMA4
Assay: Agonist activity at mouse GPR120 expressed in U2OS cells assessed as induction of beta-arrestin2 recruitment after 90 mins by luminescence assay
Measurement: EC50 = 5300 nM (pChEMBL 5.28000020980835)
Descriptors: MW 351.87, ALogP 4.6, HBA 3, HBD 1, PSA 49.77, QED 0.86
Compound (SMILES): CCC(=O)NC1CCc2c(-c3cc4c(cc3F)N(C)C(=O)CC4)cncc21
InChIKey: RXJOFZNMHGDABT-UHFFFAOYSA-N
Target: Cytochrome P450 11B2, mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: P19099
Assay: Cellular Enzyme Assay: The expression plasmids contains the ORF for either human/cyno CYP11B1 or CYP11B2 under the control of a suitable promoter (CMV-promoter) and a suitable resistance marker (neomycin). Using standard techniques the expression plasmid is transfected into G-402 cells and these cells are then selected for expressing the given resistance markers. Individual cell-clones are then selected and assessed for displaying the desired enzymatic activity using 11-Deoxycorticosterone (Cyp11B2) or 11-Deoxycortisol (Cyp11B1) as a substrate. G-402 cells expressing CYP11 constructs were established as described above and maintained in McCoy's 5a Medium Modified, ATCC Catalog No. 30-2007 containing 10% FCS and 400 μg/ml G418 (Geneticin) at 37° C. under an atmosphere of 5% CO2/95% air. Cellular enzyme assays were performed in DMEM/F12 medium containing 2.5% charcoal treated FCS and appropriate concentration of substrate (0.3-10 μM 11-Deoxycorticosterone, 11-Deoxycortisol or Corticosterone). For assaying enzymatic activity, cells were plated onto 96 well plates and incubated for 16 h. An aliquot of the supernatant is then transferred and analyzed for the concentration of the expected product (Aldosterone for CYP11B2; Cortisol for CYP11B1). The concentrations of these steroids can be determined using HTRF assays from CisBio analyzing either Aldosterone or Cortisol.
Measurement: EC50 = 1377.9 nM (pChEMBL 5.860000133514404)
Descriptors: MW 367.42, ALogP 3.31, HBA 3, HBD 1, PSA 62.3, QED 0.91
Compound (SMILES): NCc1ccnc(NC(=O)[C@@H]2CC[C@@H]3CN2C(=O)N3OS(=O)(=O)O)c1
InChIKey: KILBLDFFQRBEMS-ZJUUUORDSA-N
Target: Beta-lactamase (SINGLE PROTEIN), Klebsiella pneumoniae
UniProt: Q93LQ9
Assay: Spectrophotometric Assay: Enzyme activities were measured in the presence of the test inhibitor in spectrophotometric assay.
Measurement: IC50 = 1300 nM (pChEMBL 5.889999866485596)
Descriptors: MW 371.38, ALogP -0.52, HBA 7, HBD 3, PSA 155.16, QED 0.58
Compound (SMILES): COc1ccc(C(=O)C2CCN(C3CCN(CC(=O)Nc4nnnn4C)C3=O)CC2)cc1
InChIKey: KAECKBGXSOKYSN-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase tankyrase-1 (SINGLE PROTEIN), Homo sapiens
UniProt: O95271
Assay: Biochemical Assay: The autoparsylation activity of the TNKS 1/2 or PARP1/2 enzymes was measured by the liquid chromatography-mass spectrometry (LC/MS) detection of nicotinamide as readout. Compound activity in inhibiting the TNKS and PARP autoparsylation was evaluated by IC50 measurements. In the compound screening assays, the reaction is composed of 5 uL of compound in 8-point serial dilutions with concentrations ranging from 0.0086 to 18.75 uM, 20 nM of purified enzyme, and 250 uM of p-NAD+ in the 1x Assay Buffer. After 60 min incubation at room temperature, the reactions were quenched by the addition of 10 uL of 5x quenching solution (20% formic acid and 500 nM [d]-nicotinamide in water). For the background control wells, 10 uL of the 5x quenching solution per well was added prior to the addition of p-NAD+. The % Inhibition was calculated as: (Control-Sample)/(Control-Background)*100. "Control" is the average value of 8 wells without compound; and "Background" is the average of 8 wells mixed with 5x quenching solution measured prior to initiation of the reaction.
Measurement: IC50 = 3080 nM (pChEMBL 5.510000228881836)
Descriptors: MW 441.49, ALogP 0.35, HBA 9, HBD 1, PSA 122.55, QED 0.61
Compound (SMILES): CS(=O)(=O)N1CCn2c(CCc3ccccc3)c(C(F)(F)F)c3cccc(c32)C1
InChIKey: VCZPAUZUTMEULW-UHFFFAOYSA-N
Target: Endothelial lipase (SINGLE PROTEIN), Mus musculus
UniProt: Q9WVG5
Assay: Endothelial Lipase Assay: To assay for cell surface lipase activity, cells expressing human endothelial lipase (EL) or LPL were plated in CellBIND® 384-well plates (Corning, Lowell, Mass.) in 25 μL serum free medium at a density of 2000 cells/well. After 18-24 hours incubation at 37° C., the medium was removed and replaced with 15 μL assay buffer [Hank's Buffered Saline Solution with 25 mM HEPES pH 7.2] and 15 μL PLA1 substrate for a final concentration of 10 μM using a Multidrop reagent dispenser. Fluorescence signal was monitored for 30 min at 37° C. on a Safire II plate reader in kinetic mode (60 cycles, kinetic interval: 30 seconds) with an excitation wavelength of 490 nm and an emission wavelength of 515 nm.
Measurement: IC50 = 340 nM (pChEMBL 6.46999979019165)
Descriptors: MW 422.47, ALogP 4.22, HBA 3, HBD 0, PSA 42.31, QED 0.63
Compound (SMILES): CN[C@@H](C)C(=O)N[C@@H]1C(=O)N(C(C)C)c2ccccc2OC12CCOCC2
InChIKey: WZDTUXLPMXNSTO-WMLDXEAASA-N
Target: E3 ubiquitin-protein ligase XIAP (SINGLE PROTEIN), Homo sapiens
UniProt: P98170
Assay: TR-FRET Assay: Ten nanomolar of 6x Histidine-tagged BIR2 domain, corresponding to amino acids 124-240 of XIAP, or BIR3 domain, corresponding to amino acids 241-356 of XIAP, was mixed with 20 nM of the peptide AVPIAQKSEK-(8-biotin)-OH 1:2 TFA, in the presence of 50 mM Tris-C1, pH 7.5, 100 mM NaCl, 1 mM dithiothreitol (DTT) and 0.1 mg/mL bovine serum albumin (BSA). Following a 45 min. incubation at 37° C., Europium-Streptavidin and Allophycocyanin conjugated anti-Histidine antibody were added to a final concentration of 1.5 nM and 15 nM, respectively. Time-resolved fluorescence resonance energy transfer (TR-FRET) signals were measured 1 hour later at room temperature. Test compound potency was assessed at 10 serially diluted concentrations.
Measurement: IC50 = 54800 nM (pChEMBL 4.260000228881836)
Descriptors: MW 375.47, ALogP 1.46, HBA 5, HBD 2, PSA 79.9, QED 0.83
Compound (SMILES): Nc1ccccc1NC(=O)CCCCCNC(=O)c1ccc2c(c1)OCCO2
InChIKey: BKQDTCGCWHSJTC-UHFFFAOYSA-N
Target: Histone deacetylase 3 (SINGLE PROTEIN), Homo sapiens
UniProt: O15379
Assay: HDAC enzyme inhibitionAssay: HDAC enzyme inhibition assays were performed using purified HDACs 1-10 essentially as described in Beckers et al., 2007, Int. J. Cancer., 121:1138-48 and Perez-Balado et al., 2007, J. Med. Chem., 50:2497-2505. Inhibition assays using nuclear extract were performed essentially as described in Herman et al., 2006, Nat. Chem. Biol., 2:551-558. Briefly, the purified HDACs or nuclear extract were incubated with an acetylated substrate in the absence of the compound to be assayed and with increasing concentrations of the compound. The rate of substrate deacetylation was measured under each condition, and half-maximal inhibitory concentration with regard to each HDAC was determined by standard means.
Measurement: IC50 = 585 nM (pChEMBL 6.230000019073486)
Descriptors: MW 383.45, ALogP 2.97, HBA 5, HBD 3, PSA 102.68, QED 0.48
Compound (SMILES): COc1cc(C#N)cnc1C(=O)Nc1ccc(F)c([C@]2(CF)C[C@@H](C(F)(F)F)OC(N)=N2)c1
InChIKey: GSULBDRDQOJUHA-HNAYVOBHSA-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 = 32.2 nM (pChEMBL 7.489999771118164)
Descriptors: MW 469.37, ALogP 3.18, HBA 7, HBD 2, PSA 122.62, QED 0.65
Compound (SMILES): N#CCC1(n2cc(C(N)=O)c(Nc3ccc(F)cc3)n2)CCN(c2cccc(F)c2)CC1
InChIKey: REILYNPBUVVEBG-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: HTRF Biochemical Assay: The reactions employed a common peptide substrate, LCB-EQEDEPEGDYFEWLW-NH2 (in-house). The basic assay protocol is as follows: First, 250 nL of diluted compounds in DMSO were dispensed into the wells of a dry 384-well Black plate (Greiner #781076) using a Labcyte Echo 555 acoustic dispenser. Subsequent reagent additions employed an Agilent Bravo. Next, 18 uL of 1.11x enzyme and 1.11x substrate in 1x assay buffer (Invitrogen kinase buffer # PV3189, 2 mM DTT, 0.05% BSA) were added to the wells and shaken and then preincubated for 30 minutes at room temperature to allow compound binding to equilibrate. After equilibration, 2 uL of 10xATP in 1x assay buffer was added to initiate the kinase reaction and the plates were shaken and then incubated at room temperature for 120 minutes. At the end of the incubation, 20 uL of 2x stop buffer (streptavidin-Dylight 650 (Thermo #84547B/100 mL), Eu-tagged pY20 antibody (Perkin Elmer #AD0067), EDTA, HEPES, and Triton) was added to quench the reaction.
Measurement: IC50 = 24 nM (pChEMBL 7.619999885559082)
Descriptors: MW 436.47, ALogP 3.91, HBA 6, HBD 2, PSA 99.97, QED 0.61
Compound (SMILES): N#CCC1(n2cc(-c3ccnc4[nH]ccc34)cn2)CN(C2CCN(C(=O)Nc3ccc(C#N)cc3F)CC2)C1
InChIKey: BSLCYAXDLCRAAM-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Inhibition of human N-terminal His-tagged JAK2 catalytic domain (828 to 1132 residues) expressed in baculovirus infected insect cells assessed as decrease in phosphorylation of biotinylated peptide substrate after 1 hr in presence of 1 mM ATP by HTRF method
Measurement: IC50 > 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 523.58, ALogP 4.06, HBA 7, HBD 2, PSA 129.66, QED 0.41
Compound (SMILES): COC(=O)N[C@H](C(=O)N1CCC[C@H]1c1nc2cc([C@H]3CC[C@H](c4cc5nc([C@@H]6CCCN6C(=O)[C@@H](NC(=O)OC)C(C)C)[nH]c5cc4F)N3c3cc(F)c(N4CCCCC4)c(F)c3)c(F)cc2[nH]1)C(C)C
InChIKey: IELDPIIQUCACBU-CNGYLHDYSA-N
Target: Nonstructural protein 5A (SINGLE PROTEIN), Hepatitis C virus
UniProt: Q5L478
Assay: Inhibition of Hepatitis C virus genotype 1a H77 replicon NS5A Y93H mutant transfected in human HuH7 cells assessed as decrease in replication of viral replicon after 4 days by luciferase reporter gene assay
Measurement: EC50 = 0.08 nM (pChEMBL 10.100000381469727)
Descriptors: MW 987.11, ALogP 9.16, HBA 10, HBD 4, PSA 181.12, QED 0.09
Compound (SMILES): CC1(C2CC(=O)N(CCOc3ccc(F)c(C#N)c3)C2)CCS(=O)(=O)CC1
InChIKey: SYEMWCXBADSSGE-UHFFFAOYSA-N
Target: Platelet-activating factor acetylhydrolase (SINGLE PROTEIN), Homo sapiens
UniProt: Q13093
Assay: Inhibition of recombinant human Lp-PLA2 pre-incubated for 30 mins before PED6 fluorogenic substrate
Measurement: IC50 = 180 nM (pChEMBL 6.75)
Descriptors: MW 394.47, ALogP 2.14, HBA 5, HBD 0, PSA 87.47, QED 0.76
Compound (SMILES): O=C(CNCC(F)(F)c1cccc(OCCCC(F)(F)F)c1)N1CCCC1
InChIKey: SXYKINBAZCLVRK-UHFFFAOYSA-N
Target: Cytochrome P450 2D6 (SINGLE PROTEIN), Homo sapiens
UniProt: P10635
Assay: Inhibition Assay: Test compound is dissolved in DMSO 500× the highest final concentration desired in the IC50 assay. 30 ml of deionized water is pre-warmed to 37° C. and all kit components are placed on ice. For each well of column 1, 149.4 μL of NADPH-Cofactor Mix, (187.5 μl of Cofactors, 150 μl of G6PDH, 100 μl of Control Protein and 14.56 ml of 37° C. water) are added. In each well from Column 2 to 12, 100 μl of Cofactor/DMSO mix (40 μL DMSO in 9.96 ml of NADPH-Cofactor Mix) are added. To each well of column 1, 0.6 μl of test compound or positive control are added. 50 μl from each well of column 1 are serially diluted to column 8. The extra 50 μl from column 8 are discarded. The plate is covered and pre-incubated at 37° C. for 10 minutes. Preparation of enzyme/substrate mix: 7.92 ml of pre-warmed deionized water, 75 μl of enzyme, 3 μl of 10 mM AMMC and 2 ml of pre-warmed buffer are mixed. After the pre-incubation time (10'), 100 μl of enzyme/substrate mix to each well from column 1 to 10 are added. The plate is incubated at 37° C. for 30 minutes. After this time, 75 μl of Stop Reagent to each well are added. For blank controls, 100 μl of enzyme/substrate mix are added to columns 11 and 12.
Measurement: IC50 > 40000 nM (pChEMBL 4.3979997634887695)
Descriptors: MW 394.38, ALogP 3.71, HBA 3, HBD 1, PSA 41.57, QED 0.51
Compound (SMILES): O=S(=O)(Nc1ncns1)c1ccc2c(c1)OCCN2c1cccc(OC(F)(F)F)c1
InChIKey: RXNZKMCVXFBVOX-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 = 138 nM (pChEMBL 6.860000133514404)
Descriptors: MW 458.44, ALogP 3.77, HBA 8, HBD 1, PSA 93.65, QED 0.62
Compound (SMILES): O=C(O)c1ccc(C(=O)N2CCC(Nc3cc(=O)[nH]c4ccc(C(c5ccc(Cl)cc5)c5ccc(Cl)cc5)cc34)CC2)cc1
InChIKey: HLXBZHWWDPFRAD-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 626.54, ALogP 7.43, HBA 4, HBD 3, PSA 102.5, QED 0.16
Compound (SMILES): CN1C(=N)N[C@](C)(c2sc3c(-c4ccc(C5CC(=O)N5)cc4)cccc3c2F)CS1(=O)=O
InChIKey: XEVRRLFIUKBQIP-XLDIYJRPSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: HTRF FRET Assay: Inhibitor compounds, prepared at 3× the desired final concentration in 1×BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO are pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1×BACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30° C. The assay is initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 μM for 4 μM for autoBACE-2) prepared in 1×BACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C. DMSO is present at 5% final concentration in the assay. Following laser excitation of sample wells at 320 nm, the fluorescence signal at 620 nm is collected for 400 ms following a 50 μs delay on a RUBYstar HTRF plate reader (BMG Labtechnologies).
Measurement: Ki = 1715 nM (pChEMBL 5.769999980926514)
Descriptors: MW 472.57, ALogP 3.28, HBA 5, HBD 3, PSA 102.36, QED 0.51