lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): COc1cc([C@H]2CC[C@@H](C)CC2)c(F)cc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: ZJGFZIHCEHNNLA-CALCHBBNSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Nav1.7 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 = 10 nM (pChEMBL 8)
Descriptors: MW 511.58, ALogP 5.22, HBA 7, HBD 1, PSA 103.43, QED 0.38
Compound (SMILES): CCN(C)C[C@@]1(C(=O)Nc2cc(C(F)(F)F)cc(C(F)(F)F)c2)CN(c2c(C)cccc2C)C(=O)[C@H]1C
InChIKey: LDFYKQPNYPZUHZ-MZNJEOGPSA-N
Target: Chemerin-like receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99788
Assay: Displacement of [125I]-chemerin from ChemR23 (unknown origin) transfected in mouse BaF3 cells after 3 hrs by scintillation counting method
Measurement: IC50 <= 30 nM (pChEMBL 7.5229997634887695)
Descriptors: MW 529.53, ALogP 5.9, HBA 3, HBD 1, PSA 52.65, QED 0.47
Compound (SMILES): NS(=O)(=O)OC[C@@H]1C[C@@H](Oc2ncnc3nc(-c4c[nH]c5ccccc45)[nH]c23)C[C@@H]1O
InChIKey: GELKIUGEMSHZAZ-RWSFTLGLSA-N
Target: NEDD8 activating enzyme (PROTEIN COMPLEX), Homo sapiens
UniProt: Q13564
Assay: Inhibition of recombinant human NAEalpha/N-terminal His-tagged NAEbeta expressed in baculovirus infected insect Sf9 cells using N-terminal FLAG tagged Nedd8 as substrate incubated for 105 mins in presence of N-terminal GST-tagged Ubc12 by FRET assay
Measurement: IC50 <= 500 nM (pChEMBL 6.301000118255615)
Descriptors: MW 444.47, ALogP 1.24, HBA 8, HBD 4, PSA 169.1, QED 0.34
Compound (SMILES): Cc1ccc(N2CC(C(=O)Nc3cc(C(F)(F)F)cc(C(F)(F)F)c3)CC2=O)c(C)c1
InChIKey: FFMRFFZDGIWPLH-UHFFFAOYSA-N
Target: Chemerin-like receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99788
Assay: Displacement of [125I]-chemerin from ChemR23 (unknown origin) transfected in mouse BaF3 cells after 3 hrs by scintillation counting method
Measurement: IC50 < 30 nM (pChEMBL 7.5229997634887695)
Descriptors: MW 444.38, ALogP 5.33, HBA 2, HBD 1, PSA 49.41, QED 0.65
Compound (SMILES): CCOc1ccc(-c2nc(-c3cccc4c3CCN4CC(N)(CO)CO)no2)cc1OCC
InChIKey: KBLICPOFCCULHE-UHFFFAOYSA-N
Target: Sphingosine 1-phosphate receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21453
Assay: [35S]-GTPgamaS Binding Assay: Selected Compounds of the Examples were evaluated at Millipore Corporation, USA, using S1P1 receptor; [35S]-GTPgamaS binding assay. A [35S]-GTPgamaS binding assay at Millipore was conducted by GPCR Profiler Custom Service Laboratory, Temecula, Calif., Millipore, Inc. to monitor dose-dependent agonist selectivity for selected Examples against the S1P1 receptors. The assay was completed with sample compounds subjected to an eight-point, four-fold dose response curve with starting concentration of 10 uM. Selectivity was determined upon initial addition of compounds followed by a 30 minute incubation at 30° C. Following compound incubation, bounded [35S]-GTPgamaS was determined by filtration and scintillation counting. Independently, selected compounds were evaluated for S1P1 and S1P3 agonistic activity. The S1P1 assay system was GTPgama-S35 binding in membranes from CHO K1 cells, expressing S1P1 human receptor. The S1P3 assay system was calcium mobilization in CHO K1 cells expression.
Measurement: EC50 = 6.8 nM (pChEMBL 8.170000076293945)
Descriptors: MW 454.53, ALogP 2.25, HBA 9, HBD 3, PSA 127.1, QED 0.42
Compound (SMILES): COCCNC1(c2ccc(NC(=O)c3nc(C#N)c[nH]3)c(C3=CCC(C)(C)CC3)c2)CCCCC1
InChIKey: CPPJUTXVYRSDQU-UHFFFAOYSA-N
Target: Macrophage colony-stimulating factor 1 receptor (SINGLE PROTEIN), Mus musculus
UniProt: P09581
Assay: Fluorescence Polarization Competition Immunoassay: An autophosphorylation, fluorescence polarization competition immunoassay was used to determine the potency for c-fms inhibition exhibited by selected compounds of Formula I. The assay was performed in black 96-well microplates (LJL BioSystems). The assay buffer used was 100 mM 4-(2-hydroxyethyl)piperazine 1-ethanesulfonic acid (HEPES), pH 7.5, 1 mM 1,4-dithio-DL-threitol (DTT), 0.01% (v/v) Tween-20. Compounds were diluted in assay buffer containing 4% dimethylsulfoxide (DMSO) just prior to the assay. To each well, 5 μL of compound were added followed by the addition of 3 μL of a mix containing 33 nM c-fms (Johnson & Johnson PRD) and 16.7 mM MgCl2 (Sigma) in assay buffer. The kinase reaction was initiated by adding 2 μL of 5 mM ATP (Sigma) in assay buffer. The final concentrations in the assay were 10 nM c-fms, 1 mM ATP, 5 mM MgCl2, 2% DMSO. Control reactions were ran in each plate: in positive and negative control wells, assay buffer (made 4% in DMSO) was substituted for the compound; in addition, positive control wells received 1.2 μL of 50 mM ethylenediaminetetraaceticacid (EDTA). The plates were incubated at room temperature for 45 min. At the end of the incubation, the reaction was quenched with 1.2 μL of 50 mM EDTA (EDTA was not added to the positive control wells at this point; see above). Following a 5-min incubation, each well received 10 μL of a 1:1:3 mixture of anti-phosphotyrosine antibody, 10×, PTK green tracer, 10× (vortexed), FP dilution buffer, respectively (all from PanVera, cat. # P2837). The plate was covered, incubated for 30 min at room temperature and the fluorescence polarization was read on the Analyst. The instrument settings were: 485 nm excitation filter; 530 nm emission filter; Z height: middle of well; G factor: 0.93. Under these conditions, the fluorescence polarization values for positive and negative controls were approximately 300 and 150, respectively, and were used to define the 100% and 0% inhibition of the c-fms reaction.
Measurement: IC50 = 2.9 nM (pChEMBL 8.539999961853027)
Descriptors: MW 475.64, ALogP 5.52, HBA 5, HBD 3, PSA 102.83, QED 0.44
Compound (SMILES): O=S1(=O)C=C(c2ccc(Br)cc2)S(=O)(=O)CCC1
InChIKey: TWIDQZOSSNZKQN-UHFFFAOYSA-N
Target: Galanin receptor type 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P47211
Assay: Displacement of [125I]human galanin from human galanin-1 receptor after 16 hrs by scintillation proximity assay
Measurement: IC50 = 260 nM (pChEMBL 6.579999923706055)
Descriptors: MW 351.24, ALogP 1.98, HBA 4, HBD 0, PSA 68.28, QED 0.78
Compound (SMILES): CCCCS(=O)(=O)Nc1ccc(NC(=O)N2Cc3ccncc3C2)cc1
InChIKey: LUWXXTONVVPGRU-UHFFFAOYSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Binding Assay : Probe 2:The assay was carried out in 18 uL low volume plates (Owens Corning) in reaction buffer (50 mM HEPES (NaOH), pH 7.5, 100 mM NaCl, 10 mM MgCl2, 1 mM DTT, 1% Glycerol) using 6.8 nM recombinant, human, C-terminally-His tagged NAMPT, 1 nM Tb-anti-His antibody (Invitrogen, Cat #PV5895), and 200 nM probe (Oregon Green 488-conjugated APO0866; A-1287128.0. Plates were covered, and reactions were carried out for 2-3 hours. Plates were read with Envision (Laser Lantha low volume protocol) after 2 to 3 hours. Excitation was carried out at 337 nm, and the ratio of emission of Oregon Green (520 nm) to terbium (492 nm) was determined and used to calculate IC50 values of test compounds.
Measurement: IC50 = 39.8 nM (pChEMBL 7.400000095367432)
Descriptors: MW 374.47, ALogP 3.17, HBA 4, HBD 2, PSA 91.4, QED 0.81
Compound (SMILES): CC1(C)OC(c2ccnc(NCCC#N)n2)=C(c2cc(Cl)cc(NS(=O)(=O)c3cc(F)ccc3F)c2F)C1=O
InChIKey: UZKDSDZTDBBBPX-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: LanthaScreen Kinase Assay: The BRAFV600E enzymatic assay was performed using a LanthaScreen kinase assay kit purchased from Life Technologies (Grand Island, N.Y.). The assay was conducted according to the procedure provided in the assay kit. In brief, the enzyme reaction was carried out in the kinase reaction buffer containing BRAFV600E (20 ng/mL), ATP (2 uM), Fluorescein-MAP2K1 inactive substrate (0.4 uM), HEPES (50 mM, pH 7.5), 0.01% BRIJ-35, MgCl2 (10 mM), and EGTA (1 mM) in the presence or absence of the tested compounds at various concentrations in a 384-well plate at room temperature (22±1° C.) for 60 minutes. The final reaction volume for each reaction was 10 ul. The reaction was stopped by addition of 10 ul of TR-FRET dilution buffer kinase supplemented with kinase quench buffer (10 mM final) and Tb-anti-pMAP2K1 (2 nM final). The plate was further incubated at room temperature for another 60 minutes, and the fluorescent signals were read on Victor 5 (Perkin Elmer) with excitation at 340 nM and emission at 495 and 520 nM. The assay signal was determined as a ratio of FRET-specific signal measured with emission filter at 520 nM to that of the signal measured with Tb-specific emission filter at 495 nM. IC50 value was calculated using appropriate programs in GraphPad Prism by plotting the logarithm of the concentration versus percent inhibition.
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 577.97, ALogP 4.92, HBA 8, HBD 2, PSA 134.07, QED 0.36
Compound (SMILES): CC(=O)N[C@H](C(=O)N[C@@H](CCCNC(=N)N)C(=O)NC1(C(N)=O)CC1)C(C)C
InChIKey: CYLASULWGHWYGQ-RYUDHWBXSA-N
Target: WD repeat-containing protein 5 (SINGLE PROTEIN), Homo sapiens
UniProt: P61964
Assay: Binding Assay: All the FP based experiments were performed in MICROFLUOR 2 Black, "U" Bottom, 96-well Microtiter Plates (ThermoSci.) and FP was measured as mP units in a microplate reader (Tecan Ultra) with excitation at 485 nm and emission at 530 nm. To dilutions of WDR5Δ23 (2.2-0 uM) in 100 ul assay buffer (0.1M Phosphate, 25 mM KCl, 0.01% Triton, pH 6.5) 20 ul of a fixed concentration of the tracer in the assay buffer was added, followed by an addition of 5 ul DMSO to give 125 ul of total volume. Each assay had two controls: blank (without protein and tracer) and tracer only (without protein). The plates were incubated on a shaker at room temperature to reach equilibrium and mP values were measured at the 3 hour time point.
Measurement: IC50 < 10000 nM (pChEMBL 5)
Descriptors: MW 397.48, ALogP -1.97, HBA 5, HBD 7, PSA 192.29, QED 0.12
Compound (SMILES): Cc1cncc(C(=O)Nc2cccc(CN3CCC(C(=O)NC(C)C)CC3)c2)c1
InChIKey: NSVBQLKUVFTMTE-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 394.52, ALogP 3.38, HBA 4, HBD 2, PSA 74.33, QED 0.79
Compound (SMILES): O=C(O)c1cccc(S(=O)(=O)N2CCc3ccc(C#Cc4ccc(C(F)(F)F)cc4)cc3C2)c1
InChIKey: QLUHOEBQXBSPDV-UHFFFAOYSA-N
Target: Aldo-keto reductase family 1 member C3 (SINGLE PROTEIN), Homo sapiens
UniProt: P42330
Assay: Inhibition of human recombinant His-tagged AKR1C3 expressed in Escherichia coli BL21 (DE3) cells using 8-Acetyl-2,3,5,6-tetrahydro-1H,4H-11-oxa-3a-aza-benzo[de]anthracen-10-one as substrate after 1 hr in presence of NADPH by fluorescence assay
Measurement: IC50 = 1310 nM (pChEMBL 5.880000114440918)
Descriptors: MW 485.48, ALogP 4.55, HBA 3, HBD 1, PSA 74.68, QED 0.55
Compound (SMILES): CO[C@H]1CC[C@]2(CC1)Cc1ccc(OCc3ccccc3)cc1C21N=C(N)N(C)O1
InChIKey: CIPLWUBAYCECKC-AHLJTMJASA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: BACE Assay: For each compound being tested, the BACE activity was monitored in a fluorescence quenching assay (FRET) using the ectodomain of BACE (aa 1-454) fused to a myc-his tag and secreted from HEK293/BACEect. cells into OptiMEM (Invitrogen) as enzyme source and a substrate peptide derived from the APP-Swedish mutation which possesses a Cy3-fluorophore at the N-terminus and a Cy5Q-quencher at the C-terminus (Cy3-SEVNLDAEFK-Cy5Q-NH2; Amersham).
Measurement: IC50 = 890 nM (pChEMBL 6.050000190734863)
Descriptors: MW 407.51, ALogP 3.74, HBA 6, HBD 1, PSA 69.31, QED 0.84
Compound (SMILES): CC(C)(O)c1cn(-c2ccc(-c3cc(F)c(CO)c(S(C)(=O)=O)c3)cc2)c(C(C)(C)c2ccccc2F)n1
InChIKey: PABVYBCMEDLXBK-UHFFFAOYSA-N
Target: Retinoic acid receptor RXR-alpha/oxysterols receptor LXR-beta (PROTEIN COMPLEX), Homo sapiens
UniProt: P19793
Assay: Displacement of [3H]-24,25-epoxycholesterol from human LXRbeta/RXRalpha expressed in baculovirus infected Sf9 cells by scintillation proximity analysis
Measurement: Ki = 14 nM (pChEMBL 7.849999904632568)
Descriptors: MW 540.63, ALogP 5.27, HBA 6, HBD 2, PSA 92.42, QED 0.33
Compound (SMILES): CC(NC(=O)c1cc(NC(=O)c2cc(-c3ccccn3)c(F)cc2Cl)n(-c2ccccc2)n1)c1nncn1C
InChIKey: SWMWLDXNGDRXPZ-UHFFFAOYSA-N
Target: High affinity nerve growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P04629
Assay: Cell Based Assay: TRKA co-expressed with p75: The assays are based upon DiscoveRx's proprietary Enzyme Fragment Complementation (EFC) technology. In the case of the TRK cell lines, the enzyme acceptor (EA) protein is fused to a SH2 protein and the TRK receptor of interest has been tagged with a Prolink tag. Cryopreserved PathHunter cells are used from either in-house produced batches or bulk batches bought directly from DiscoveRx. Cryopreserved cells are resuscitated, spun 1000 rpm for 4 min to remove freezing media, and resuspended in MEM+0.5% horse serum (both Invitrogen) to 5e^5 cells/ml. The cells are then plated using a Multidrop into Greiner white tissue culture treated plates at 20 μl/well and incubated for 24 h at 37 °C., 5% CO2, high humidity. On the day of the assay, the cell plates are allowed to cool to room temperature for 30 min prior to the assay. 4 mM stock solutions of test compounds are prepared and serially diluted in 100% DMSO. A standard curve using the compound of Example 135, WO2005/116035 at a top concentration of 150 μM is also prepared on each test plate. High percentage effect (HPE) is defined by 150 μM of the compound of Example 135, WO2005/116035 and 0% effect (ZPE) is defined by 100% DMSO. Plates containing 1 μl of serially diluted compound, standard and HPE/ZPE are diluted 1/66 in assay buffer (PBS minus Ca2+, minus Mg2+ with 0.05% pluronic F127) using a Wellmate. Using a Platemate Plus, 5 μl of 1/66 diluted test compounds is then transferred to the cell plate and allowed to reach equilibrium by incubating for 30 min at room temperature before addition of agonist stimulus: 10 μl/well of 2 nM (0.571 nM FAC) of the cognate neurotrophin (Peprotech) diluted in agonist buffer (HBSS with 0.25% BSA). Final assay concentration of the test compounds is 8.66 μM, (the compound of Example 135, WO2005/116035 FAC is 0.325 μM). The plates are left at room temperature for a further 2 hours before addition of 10 μl of the DiscoveRx PathHunter detection reagent (made up by adding 1 part Galacton Star, 5 parts Emerald II and 19 parts Cell Assay Buffer as per the manufacturer's instructions). After reagent addition, plates are covered and incubated at room temperature for 60 minutes. Luminescence signal is read using an Envision.
Measurement: IC50 = 298 nM (pChEMBL 6.53000020980835)
Descriptors: MW 544.98, ALogP 4.6, HBA 8, HBD 2, PSA 119.62, QED 0.31
Compound (SMILES): Cl.N=C(N)N/N=C/c1ccc(-c2ccc(/C=N/Nc3ccccc3Br)cc2)cc1
InChIKey: BBGOSHDIHDXIDF-RXQQCVBLSA-N
Target: Voltage-gated inwardly rectifying potassium channel KCNH2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q12809
Assay: Displacement of [3H]-dofetilide from human ERG expressed in HEK293 cell membranes
Measurement: IC50 = 440 nM (pChEMBL 6.360000133514404)
Descriptors: MW 435.33, ALogP 4.38, HBA 4, HBD 4, PSA 98.65, QED 0.26
Compound (SMILES): Cc1ccc(NC(=O)c2cccc(C(F)(F)F)c2)cc1-c1cnc(OC(C)C)c(N2CCOCC2)c1
InChIKey: RINDPRPVBOGYGN-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: In Vitro Enzyme Assay: Final assay conditions were 4 pM cRaf, 3 uM ATP, 10 nM biotin tagged MEK1 kinase dead protein substrate. Reactions were performed in Greiner384 well plates, catalog #784075. Reaction buffer was 50 mM Tris, pH 7.5, 50 mM NaCl, 10 mM MgCl2, 0.01% Tween-20, 0.05% BSA and 1 mM DTT. 5 ul of 2×cRaf plus MEK1 were dispensed into plates containing 0.25 ul of compound in 100% DMSO and incubated at room temperature for 30 minutes. Reactions were started by addition of 5 ul of 2×ATP. Reactions were run for two hours and terminated by addition of 10 ul of 2× antibody and Alphascreen bead mixture in Stop Buffer. Stop Buffer was 50 mM Tris, pH 7.5, 0.005% Tween-20, 25 mM EDTA.
Measurement: IC50 = 4.173 nM (pChEMBL 8.380000114440918)
Descriptors: MW 499.53, ALogP 5.95, HBA 5, HBD 1, PSA 63.69, QED 0.45
Compound (SMILES): Fc1ccc(CNc2n[nH]c3ccc(-c4cn(Cc5ccccc5)nn4)cc23)c(F)c1
InChIKey: XZAMYBUMHZTQDY-UHFFFAOYSA-N
Target: Cell division cycle 7-related protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: O00311
Assay: Inhibition Assay: 11 data point. Cdc7 kinase assays were carried out in 25 mM HEPES, pH 7.5, 1 mM DTT, 10 mM MgCl2, 100 μM Na3VO4, and 0.075 mg/ml Triton X-100 using 12 ng baculovirus-expressed Cdc7/Dbf4 and 2 μM Jerini peptide substrate A-A11 (biotin-C6linker-TPSDSLIYDDGLS) (SEQ ID NO:5). Reactions were initiated with λ-[33P]-ATP (1 μM, 20 mCi/μmol) and quenched after 1 hour with 5 volumes of stop buffer (50 mM EDTA, 2 M NaCl). The reactions were incubated for 30 minutes on streptavidin-coated plates, washed, and quantified using a TopCount scintillation plate reader (Packard). IC50 values are determined via non-linear regression fitting of the data. Ki values are generated assuming ATP-competitive (equilibrium) inhibition and using the experimentally determined apparent ATP Km of Cdc7 (0.7 μM).
Measurement: Ki = 37 nM (pChEMBL 7.429999828338623)
Descriptors: MW 416.44, ALogP 4.76, HBA 5, HBD 2, PSA 71.42, QED 0.42
Compound (SMILES): CS(=O)(=O)c1ccc(Nc2nccc(-c3ccc(Oc4ccccc4N)cc3)n2)cc1
InChIKey: IORGYFRRDJTHBE-UHFFFAOYSA-N
Target: Inhibitor of nuclear factor kappa-B kinase subunit beta (SINGLE PROTEIN), Homo sapiens
UniProt: O14920
Assay: Inhibition of IKKbeta (unknown origin)
Measurement: IC50 = 2700 nM (pChEMBL 5.570000171661377)
Descriptors: MW 432.51, ALogP 4.67, HBA 7, HBD 2, PSA 107.2, QED 0.43
Compound (SMILES): CNC(=O)c1cccc(Nc2ncnc(N3CCc4cc(C(F)(F)F)ccc4C3)n2)c1
InChIKey: HTNJKCJFVBYIGH-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Cell-Based Assay: Compounds were tested on human Nav1.7 (hNav1.7) expressed stably in 293 cells, which have no endogenous sodium channels. Two forms of whole-cell patch-clamp assays were used. In both, peak current through Nav1.7 was evoked by a depolarizing voltage pulse, and currents were monitored as a function of time in response to exposure to several different concentrations of drug. Automated electrophysiology uses the PatchXpress 7000A system. Up to sixteen cells at a time were voltage-clamped in parallel, and the steady-state voltage dependence of inactivation was determined experimentally for each cell. For each cell, fractional inactivation was set at 20%, and peak current through Nav1.7 as a function of time was measured in response to increasing concentrations of test compound. Manual electrophysiology refers to conventional whole-cell patch-clamping (Hamill et al., 1981). After formation of the whole-cell clamp, cells were lifted off the bottom of the dish with the patch pipette and positioned directly in front of an array of glass microperfusion tubes each with internal diameter about 250 microns, with individual compound concentrations flowing from each tube. Current as a function of time in response to increasing drug concentrations was measured with Nav1.7 fully in the resting state (holding voltage set to −140 mV) and on the same cells with the holding voltage set to produce about 20% inactivation. Compounds were not applied for a prespecified time, but for as long as required to achieve a stable level of block.
Measurement: IC50 = 8600 nM (pChEMBL 5.070000171661377)
Descriptors: MW 428.42, ALogP 3.56, HBA 6, HBD 2, PSA 83.04, QED 0.66
Compound (SMILES): O=c1[nH]cccc1S(=O)(=O)CC[C@@H]1OCC[C@@]2(S(=O)(=O)c3ccc(Cl)cc3)c3c(F)ccc(F)c3OCC12
InChIKey: DSRJABQQFABIIF-NHHFAVJZSA-N
Target: Amyloid-beta precursor protein (SINGLE PROTEIN), Homo sapiens
UniProt: P05067
Assay: Electrochemiluminescence (ECL)-based Immunoassay: To measure γ-secretase activity, membranes were incubated at 37° C. for 1 h in 50 μL of buffer containing 20 mM Hepes (pH 7.0) and 2 mM EDTA. At the end of the incubation, Aβ 40 and Aβ 42 were measured using an electrochemiluminescence (ECL)-based immunoassay. Aβ 40 was identified with antibody pairs TAG-G2-10 and biotin-W02, while Aβ 42 was identified with TAG-G2-11 and biotin-4G8. The ECL signal was measured using an ECL-M8 instrument (IGEN International, Inc.) according to the manufacturer's instructions.
Measurement: IC50 = 2.8 nM (pChEMBL 8.550000190734863)
Descriptors: MW 586.03, ALogP 3.64, HBA 7, HBD 1, PSA 119.6, QED 0.47
Compound (SMILES): Fc1ccc(-c2n[nH]c3ccc(/C=C/c4ccncc4)cc23)cc1
InChIKey: BBIYWBMKGNBJRZ-OWOJBTEDSA-N
Target: Mitogen-activated protein kinase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: P45984
Assay: Inhibition of His6-tagged JNK2 (unknown origin) assessed as reduction in c-Jun phosphorylation using GST-c-Jun (1 to 79 residues) and [gamma32P]ATP incubated for 1 hr by scintillation counting assay
Measurement: IC50 <= 10000 nM (pChEMBL 5)
Descriptors: MW 315.35, ALogP 4.93, HBA 2, HBD 1, PSA 41.57, QED 0.58
Compound (SMILES): CCCCn1c(=O)c(C(=O)Nc2ccc(C(=O)OCCC)cc2)c(O)c2ccccc21
InChIKey: GZKCRVUTBBOLCK-UHFFFAOYSA-N
Target: Protein smoothened (SINGLE PROTEIN), Mus musculus
UniProt: P56726
Assay: Activity at Smo in mouse C3H10T1/2 cells assessed as induction of cell differentiation into osteoblast incubated for 6 days by alkaline phosphatase assay
Measurement: EC50 = 800 nM (pChEMBL 6.099999904632568)
Descriptors: MW 422.48, ALogP 4.33, HBA 6, HBD 2, PSA 97.63, QED 0.53
Compound (SMILES): C[C@H](Nc1ncc(F)c(N2C(=O)OC[C@@H]2[C@@H](C)O)n1)c1ccc(OCC(F)(F)F)c(F)c1
InChIKey: WVFOOPQGQMFLTC-IMSIIYSGSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Fluorescence Biochemical Assay: The IDH1 (R132H) mutant catalyzes the reduced form of NADP+ (NADPH) and α-ketoglutarate (α-KG) to form nicotinamide adenine dinucleotide phosphate (NADP+) and R (−)-2-hydroxyglutarate (2HG). The reaction can be monitored kinetically by following the oxidation of NADPH to NADP+ which is measured using fluorescence, excitation at 355 nm and emission at 530 nm. Reactions were monitored using the Perkin-Elmer Envision, Model 2101. More specifically, the biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Cat. No. 781076) using a final reaction volume of 20 μL and the following assay buffer conditions: 50 mM HEPES pH 7.5, 10 mM MgCl2, 1 mM DTT, 0.02% BSA, 0.02% Tween-20, 10 μM NADPH and 100 μM α-KG. The final reaction mixture contained 2.5% DMSO and test compounds with concentrations ranging 0.0000008-25 μM. The IDH1 (R132H) enzyme was used at a final concentration of 10 nM.
Measurement: IC50 = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 462.38, ALogP 3.58, HBA 7, HBD 2, PSA 96.81, QED 0.61
Compound (SMILES): NC(=O)C12CC3CC(C1)C(NC(=O)c1cccc(NS(=O)(=O)c4ccccc4)c1)C(C3)C2
InChIKey: ZCJWATXWMDTBMO-UHFFFAOYSA-N
Target: 11-beta-hydroxysteroid dehydrogenase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P28845
Assay: HTRF Assay : The inhibiting activity of 11β-HSD1 derived from microsomal fractions was measured using the HTRF assay (62CO2PEB, Cisbio). Different concentrations of compounds were added to 96-well plates, followed by the addition of TE buffer (20 mM Tris buffer and 5 mM EDTA, pH 6.0) containing 200 μM NADPH (N1630, Sigma) and 160 nM cortisone (C2755, Sigma), The reactions were initiated by the addition of human microsomal fractions (M0317, Sigma), and were allowed to incubate for 2 h at 37° C. Europium (Eu3+) cryptate and XL665-conjugated cortisol were then added to each well and incubated for an additional 2 h at room temperature.
Measurement: IC50 = 42.7 nM (pChEMBL 7.369999885559082)
Descriptors: MW 453.56, ALogP 2.9, HBA 4, HBD 3, PSA 118.36, QED 0.62
Compound (SMILES): Cl.Clc1ccc2sc(CCCCN3CCC4CCCCC4C3)nc2c1
InChIKey: LLHIQLAPTLWBMG-UHFFFAOYSA-N
Target: D(2) dopamine receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P14416
Assay: Displacement of [3H]-N-methylspiperone from human dopamine D2 receptor by liquid scintillation counting method
Measurement: Ki = 1087 nM (pChEMBL 5.960000038146973)
Descriptors: MW 362.97, ALogP 5.78, HBA 3, HBD 0, PSA 16.13, QED 0.63
Compound (SMILES): C[C@]12CC(O)CC1CC[C@@H]1[C@@H]2CC[C@]2(C)C(c3cncc(F)c3)=CC[C@@H]12
InChIKey: ODFCIEVQPDCQMI-JJWXIMJISA-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 355.5, ALogP 5.23, HBA 2, HBD 1, PSA 33.12, QED 0.75
Compound (SMILES): Cn1cc(-c2ccc(F)c(C(F)(F)F)c2)nc1C1CCN(c2ncnc(N)c2-c2ccc(CN)cc2)CC1
InChIKey: FJJLUWKUJSZKAK-UHFFFAOYSA-N
Target: Ribosomal protein S6 kinase beta-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23443
Assay: P70S6K Enzyme Assay: P70S6K inhibitor compounds are diluted and plated in 96 well plates. A reaction mixture including the following components is then added to the compound plate to initiate the enzyme reaction; P70S6K (3 nM, T412E mutant, Millipore) is mixed with 24 uM ATP in an assay buffer containing 100 mM Hepes (pH 7.5), 5 mM MgCl2, 1 mM DTT, 0.015% Brij and 1 uM of the substrate peptide FITC-AHA-AKRRRLSSLRA-OH (derived from the S6 ribosomal protein sequence, FITC=fluorescein isothiocyanate, AHA=6-aminohexanoic acid). The reaction is incubated for 90 min at 25.degree. C., before the addition of 10 mM EDTA to stop the reaction. The proportion of substrate and product (phosphorylated) peptide is analysed on a Caliper Life Sciences Lab Chip 3000, using a pressure of -1.4 psi, and upstream and downstream voltages of -3000 and -700 respectively. Product peaks are resolved before substrate peaks on the resulting chromatograms.
Measurement: IC50 = 4.2 nM (pChEMBL 8.380000114440918)
Descriptors: MW 525.55, ALogP 5.13, HBA 7, HBD 2, PSA 98.88, QED 0.35
Compound (SMILES): Cn1cc(-c2cc(S(C)(=O)=O)ccc2OCCN2CCNC2=O)c2cc[nH]c2c1=O
InChIKey: JOOXTAGDTZFBTP-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 = 69.3 nM (pChEMBL 7.159999847412109)
Descriptors: MW 430.49, ALogP 1.34, HBA 6, HBD 2, PSA 113.5, QED 0.61
Compound (SMILES): COc1cc2ccc3c(c2cc1OC)C[C@H](N)[C@@H](c1cc(F)c(F)cc1F)O3
InChIKey: NXPAPTOHRVIGJL-LAUBAEHRSA-N
Target: Dipeptidyl peptidase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P27487
Assay: Inhibition of DPP4 (unknown origin) expressed in Sf9 cells using Gly-Pro-AMC substrate
Measurement: IC50 = 2.06 nM (pChEMBL 8.6899995803833)
Descriptors: MW 389.37, ALogP 4.28, HBA 4, HBD 1, PSA 53.71, QED 0.68
Compound (SMILES): CC(CS(C)(=O)=O)(NC(=O)c1cc(OCC(F)(F)F)c(C2CC2)cn1)C(N)=O
InChIKey: JONIPRHLQFCBNV-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1×HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37° C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30° C. for 30 min. Compounds were added to a final assay volume of 100 μl and incubated for 30 min at 30° C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 μl lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN3) and 50 μl detection solutions (20 μM mAb Alexa700-cAMP 1:1, and 48 μM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated as follows: FRET=T730-Alexa730-P(T645-B645) with P=Ru730-B730/Ru645-B645, where T730 is the test well measured at 730 nM, T645 is the test well measured at 645 nm, B730 and B645 are the buffer controls at 730 nm and 645 nm, respectively. cAMP content is determined from the function of a standard curve spanning from 10 μM to 0.13 nM cAMP.
Measurement: EC50 = 887.9 nM (pChEMBL 6.050000190734863)
Descriptors: MW 423.41, ALogP 0.92, HBA 6, HBD 2, PSA 128.45, QED 0.64
Compound (SMILES): COc1ccc(-c2ccc(Cl)c(F)c2)cc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: OZQCJAZOZLTLIV-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Nav1.7 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 = 33 nM (pChEMBL 7.480000019073486)
Descriptors: MW 525.95, ALogP 5.25, HBA 7, HBD 1, PSA 103.43, QED 0.32
Compound (SMILES): CC(C)C[C@H](NC(=O)[C@H](CCCNC(=N)N)NC(=O)[C@H](CCC(=O)O)NC(=O)[C@H](Cc1ccccc1)NC(=O)[C@H](Cc1ccc(O)cc1)NC(=O)CNC(=O)[C@H](Cc1ccccc1)NC(=O)[C@@H](NC(=O)[C@H](Cc1ccccc1)NC(=O)[C@@H](NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(N)=O)NC(=O)CCC(=O)NCCCOCCOCCOCCCNC(=O)CCCC[C@@H]1SC[C@@H]2NC(=O)N[C@@H]21)C(C)C)C(C)C)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(N)=O
InChIKey: NVHFWOAUHDEWNO-GKRFZEPPSA-N
Target: Tissue factor pathway inhibitor (SINGLE PROTEIN), Homo sapiens
UniProt: P10646
Assay: Inhibition of C-terminus 10x-His-taged human TFPI-1 (29 to 282 residues) assessed as inhibition of factor 10a using S-2222 substrate incubated for 115 mins by micro-plate reader based assay
Measurement: EC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 2745.3
Compound (SMILES): O=C(NCc1ccc(Cl)cc1Cl)N1CCC12CN(S(=O)(=O)c1ccc(Cl)cc1)C2
InChIKey: CLMMUGZVUGYNFW-UHFFFAOYSA-N
Target: Bifunctional epoxide hydrolase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34913
Assay: Enzyme Assay: Test compounds (1.6 mM stock in DMSO) were diluted 3 fold in series in DMSO and 0.8 microliters per well were added into 384-well NBS microplates (Corning). Resorufin epoxide substrate (20 uM stock in DMSO) was diluted to 5 μM with Assay Buffer (25 mM bis-Tris-HCl, pH 7.0, 1 mM DTT and 0.2 mg/ml BSA) and 8 microliters per well were added to the microplates. Thirty two microliters per well of 3.6 nM soluble epoxide hydrolase in Assay Buffer was then added. The samples were incubated at room temperature and assay signals were monitored by reading excitation at 530 nm and emission fluorescence at 590 nm on a PlateVision (Zeiss) reader every 2 minutes for 8 times.
Measurement: IC50 = 321 nM (pChEMBL 6.489999771118164)
Descriptors: MW 474.8, ALogP 4.01, HBA 3, HBD 1, PSA 69.72, QED 0.73
Compound (SMILES): Cn1c(SCCCN2CC[C@]3(C[C@@H]3c3ccc(C(F)(F)F)cc3)C2)nnc1-c1ccc(-c2ncco2)cc1
InChIKey: NIFGQHGFOUMQJN-KCWPFWIISA-N
Target: D(3) dopamine receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35462
Assay: Displacement of [3H]-spiperone from human dopamine D3 receptor expressed in CHO-K1 cell membranes after 90 mins by liquid scintillation counting
Measurement: Ki = 0.06761 nM (pChEMBL 10.170000076293945)
Descriptors: MW 539.63, ALogP 6.52, HBA 7, HBD 0, PSA 59.98, QED 0.19
Compound (SMILES): Cl.O=C(O)CCc1ccc(OCc2ccc(CN3CCC4(CC3)CC(Cc3ccc(F)cc3)CCO4)cc2)cc1
InChIKey: XSYVABUHWWDTQM-UHFFFAOYSA-N
Target: Free fatty acid receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5NUL3
Assay: Agonist activity at human FFA4 receptor expressed in CHO cells assessed as induction of Ca2+ flux by fluo-8 dye based FLIPR assay
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 531.67, ALogP 6.43, HBA 4, HBD 1, PSA 59, QED 0.33
Compound (SMILES): CC(c1cccc(NC(=O)c2cccc(C#N)c2)c1)N1CCC(C(=O)NC2CCCCC2)CC1
InChIKey: SQPKYQMIDXBRBH-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 78 nM (pChEMBL 7.110000133514404)
Descriptors: MW 458.61, ALogP 5.03, HBA 4, HBD 2, PSA 85.23, QED 0.64
Compound (SMILES): CCOc1cc2ncnc(Nc3ccc4c(cnn4Cc4ccccc4)c3)c2cc1NC(=O)/C=C/CN1C[C@@H](C)N[C@@H](C)C1
InChIKey: AQEJOYTVPDGMEW-RRSRKQOQSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Irreversible inhibition of GST-tagged ERBB1 (unknown origin) (Met-668 to Ala-1211 residues) expressed in baculovirus infected Sf9 insect cells assessed as reduction in Glu/Tyr copolymer phosphorylation after 6 mins by ELISA
Measurement: IC50 = 17 nM (pChEMBL 7.769999980926514)
Descriptors: MW 590.73, ALogP 5.35, HBA 9, HBD 3, PSA 109.23, QED 0.19
Compound (SMILES): CC1C(c2ccccc2)CCCN1C(=O)c1ccc(N2CC(F)(F)C2)c(OCC2CC2)n1
InChIKey: YLUVLAGMBQLRKW-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1×HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37 °C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30 °C. for 30 min. Compounds were added to a final assay volume of 100 μl and incubated for 30 min at 30 °C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 μl lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN3) and 50 μl detection solutions (20 μM mAb Alexa700-cAMP 1:1, and 48 μM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated as follows: FRET=T730-Alexa730-P(T645-B645) with P=Ru730-B730/Ru645-B645, where T730 is the test well measured at 730 nM, T645 is the test well measured at 645 nm, B730 and B645 are the buffer controls at 730 nm and 645 nm, respectively. cAMP content is determined from the function of a standard curve spanning from 10 μM to 0.13 nM cAMP.
Measurement: EC50 = 40.5 nM (pChEMBL 7.389999866485596)
Descriptors: MW 441.52, ALogP 4.73, HBA 4, HBD 0, PSA 45.67, QED 0.65
Compound (SMILES): O=c1ccc2nc3ccc(OCCBr)cc3oc-2c1
InChIKey: LAWJZLFGJSXSSJ-UHFFFAOYSA-N
Target: Amyloid-beta precursor protein (SINGLE PROTEIN), Homo sapiens
UniProt: P05067
Assay: In Vitro Competitive Radioligand Binding Assay: To determine the Ki values, fixed brain tissues prepared from 15-month-old Tg2576 mice (N=5) will be incubated for 30 min with 1 uM methoxy-X34 in the presence of a serial dilution of test compounds (10-9-10-6 M, 6 concentrations/compound). Tissue sections (3 sections per reaction) will be then mounted and coverslipped. Photographs of methoxy-X34 binding images will be taken using a Nikon fluorescent microscope. The fluorescent intensity on cerebral amyloid deposits and neuritic plaques will be separately quantified using Image J software.
Measurement: Kd = 927 nM (pChEMBL 6.03000020980835)
Descriptors: MW 320.14, ALogP 3.07, HBA 4, HBD 0, PSA 52.33, QED 0.55
Compound (SMILES): O=C(NC1(c2ccccn2)CC1)c1nn(-c2ccc(C(F)(F)F)cc2Cl)c2c1CCCC/C2=C\c1ccc(Cl)s1
InChIKey: VPTHGWFPZPEPCX-BMRADRMJSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: Inverse agonist activity at human cannabinoid receptor 1 expressed in HEK293 cells assessed as increase in forskolin-induced cAMP production preincubated with cells followed by forskolin stimulation measured after 30 mins in presence of D2-labeled cAMP and Eu3+-TBP-NHS cryptate by HTRF assay
Measurement: EC50 = 8 nM (pChEMBL 8.100000381469727)
Descriptors: MW 603.5, ALogP 8.34, HBA 5, HBD 1, PSA 59.81, QED 0.23
Compound (SMILES): COc1c(OCCC(F)F)ccc2c1CCN(Cc1ccc([C@H](C)NC(C)=O)cc1)C2
InChIKey: INLMPVGVNPBILT-INIZCTEOSA-N
Target: Acetyl-CoA carboxylase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O00763
Assay: Spectrophotometric 384 Well Assay: Assay reactions are then carried out in 384-well plates, with hACC2 in an appropriate dilution and at final assay concentrations (f.c.) of 100 mM Tris (pH 7.5), 10 mM trisodium citrate, 25 mM KHCO3, 10 mM MgCl2, 0.5 mg/ml BSA, 3.75 mM reduced L-glutathione, 15 U/ml lactate dehydrogenase, 0.5 mM phosphoenolpyruvate, 15 U/ml pyruvate kinase, compounds at different concentrations at final DMSO concentrations of 1%.The enzymatic reaction is then started by addition of a mixture of NADH, acetyl Coenzyme A (both 2000 f.c.) and ATP (500 uM f.c.). The decrease of the optical density (slope S) is then determined at 25° C. at a wavelength of 340 nm over 15 minutes in a spectrophotometric reader.Each assay microtiter plate contains wells with vehicle instead of compound as controls for the non-inhibited enzyme (100% CTL; HIGH) and wells without acetyl-CoA as controls for non-specific NADH degradation (0% CTL; LOW).
Measurement: IC50 = 141 nM (pChEMBL 6.849999904632568)
Descriptors: MW 432.51, ALogP 4.48, HBA 4, HBD 1, PSA 50.8, QED 0.63
Compound (SMILES): Cc1c(-c2cccc(C(F)F)c2)c(=O)c(C(=O)NCc2ccc(S(C)(=O)=O)cn2)cn1C1CCC1
InChIKey: JRLQKETWQMVZJG-UHFFFAOYSA-N
Target: Neutrophil elastase (SINGLE PROTEIN), Homo sapiens
UniProt: P08246
Assay: Human Neutrophil Elastase Assay: Compound buffer: 100 mM Tris, 500 mM NaCl, adjusted to pH 7.5; Assay buffer: 100 mM Tris, 500 mM NaCl, adjusted to pH 7.5, containing 0.01% BSA. Test compounds were prediluted in DMSO and subsequently in compound buffer (5% DMSO final). 5 μL of these compound dilutions were mixed with 10 μl Neutrophil elastase (9 ng/ml in assay buffer) in a black 384 well OptiPlate (Perkin Elmer, Cat No.: 6007270) and incubated for 15 min at room temperature. Subsequently 10 Î¼L substrate solution in assay buffer were added (250 μM final concentration) and the plates were incubated for 60 min at room temperature. After inactivation of the enzyme, fluorescence intensities were measured at 380 nm excitation and 460 nm emission wavelengths.
Measurement: IC50 = 35 nM (pChEMBL 7.460000038146973)
Descriptors: MW 501.56, ALogP 4.21, HBA 6, HBD 1, PSA 98.13, QED 0.52
Compound (SMILES): COc1nccc2oc(-c3cnc4ccc(OCCCS(C)(=O)=NC#N)nn34)cc12
InChIKey: QPQGESIICFUNKN-UHFFFAOYSA-N
Target: MAP kinase-interacting serine/threonine-protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9BUB5
Assay: MKNK1 Kinase Assay: For the assay 50 mL of a 100 fold concentrated solution of the test compound in DMSO was pipetted into a black low volume 384 well microtiter plate (Greiner Bio-One, Frickenhausen, Germany), 2 μL of a solution of MKNK1 in aqueous assay buffer [50 mM HEPES pH 7.5, 5 mM magnesium chloride, 1.0 mM dithiothreitol, 0.005% (v/v) Nonidet-P40 (Sigma)] was added and the mixture was incubated for 15 min at 22 °C. to allow pre-binding of the test compounds to the enzyme before the start of the kinase reaction. Then the kinase reaction was started by the addition of 3 μL of a solution of adenosine-tri-phosphate (ATP, 16.7 μM=>final conc. in the 5 μL assay volume is 10 μM) and substrate (0.1 μM=>final conc. in the 5 μL assay volume is 0.06 μM) in assay buffer and the resulting mixture was incubated for a reaction time of 45 min at 22 °C. The concentration of MKNK1 was adjusted depending of the activity of the enzyme lot and was chosen appropriate to have the assay in the linear range, typical concentrations were in the range of 0.05 μg/ml. The reaction was stopped by the addition of 5 μL of a solution of TR-FRET detection reagents (5 nM streptavidine-XL665 [Cisbio Bioassays, Codolet, France] and 1 nM anti-ribosomal protein S6 (pSer236)-antibody from Invitrogen [#44921G] and 1 nM LANCE EU-W1024 labeled ProteinG [Perkin-Elmer, product no. AD0071]) in an aqueous EDTA-solution (100 mM EDTA, 0.1% (w/v) bovine serum albumin in 50 mM HEPES pH 7.5). The resulting mixture was incubated for 1 h at 22 °C. to allow the formation of complex between the phosphorylated biotinylated peptide and the detection reagents. Subsequently the amount of phosphorylated substrate was evaluated by measurement of the resonance energy transfer from the Eu-chelate to the streptavidine-XL. Therefore, the fluorescence emissions at 620 nm and 665 nm after excitation at 350 nm were measured in a TR-FRET reader, e.g. a Rubystar (BMG Labtechnologies, Offenburg, Germany) or a Viewlux (Perkin-Elmer). The ratio of the emissions at 665 nm and at 622 nm was taken as the measure for the amount of phosphorylated substrate.
Measurement: IC50 = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 426.46, ALogP 2.89, HBA 10, HBD 0, PSA 127.9, QED 0.33
Compound (SMILES): CN1CCC(C(=O)N2CC=C(c3ccc(NC(=O)N4Cc5ccccc5C4)cc3)CC2)C1
InChIKey: CYRMNXSQOKRJSP-UHFFFAOYSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Binding Assay : Probe 2:The assay was carried out in 18 uL low volume plates (Owens Corning) in reaction buffer (50 mM HEPES (NaOH), pH 7.5, 100 mM NaCl, 10 mM MgCl2, 1 mM DTT, 1% Glycerol) using 6.8 nM recombinant, human, C-terminally-His tagged NAMPT, 1 nM Tb-anti-His antibody (Invitrogen, Cat #PV5895), and 200 nM probe (Oregon Green 488-conjugated APO0866; A-1287128.0. Plates were covered, and reactions were carried out for 2-3 hours. Plates were read with Envision (Laser Lantha low volume protocol) after 2 to 3 hours. Excitation was carried out at 337 nm, and the ratio of emission of Oregon Green (520 nm) to terbium (492 nm) was determined and used to calculate IC50 values of test compounds.
Measurement: IC50 = 13.1 nM (pChEMBL 7.880000114440918)
Descriptors: MW 430.55, ALogP 3.8, HBA 3, HBD 1, PSA 55.89, QED 0.81
Compound (SMILES): Cc1cncc(C(=O)Nc2cc(CN3CCC(C(=O)NC(C)(C)C)CC3)cs2)c1
InChIKey: VPBITJKPXBAHMS-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 44 nM (pChEMBL 7.360000133514404)
Descriptors: MW 414.58, ALogP 3.83, HBA 5, HBD 2, PSA 74.33, QED 0.78