lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): C[C@H](Nc1cc(C(=O)O)nc(-c2ccc(Oc3ccc(F)cc3)cc2)n1)C(=O)O
InChIKey: LZHDSWPVMITKJX-NSHDSACASA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: FLIPR Assay: In vitro assays were performed in a recombinant cell line expressing cDNA encoding the alpha subunit (Nav1.7, SCN9a, PN1, NE) of human Nav1.7 (Accession No. NM002977). The cell line was provided by investigators at Yale University (Cummins et al, J. Neurosci. 18(23): 9607-9619 (1998)). For dominant selection of the Nav1.7-expressing clones, the expression plasmid co-expressed the neomycin resistance gene. The cell line was constructed in the human embryonic kidney cell line, HEK293, under the influence of the CMV major late promoter, and stable clones were selected using limiting dilution cloning and antibiotic selection using the neomycin analogue, G418. Recombinant beta and gamma subunits were not introduced into this cell line. Additional cell lines expressing recombinant Nav1.7 cloned from other species can also be used, alone or in combination with various beta subunits, gamma subunits or chaperones.
Measurement: IC50 > 20000 nM (pChEMBL 4.698999881744385)
Descriptors: MW 397.36, ALogP 3.66, HBA 6, HBD 3, PSA 121.64, QED 0.55
Compound (SMILES): Cc1ccc(C2=C(C(=O)NS(=O)(=O)C3CC3)C(=O)N[C@@](c3ccc(OCCCC(F)(F)F)cc3Cl)(C(F)(F)F)C2)cc1
InChIKey: KEVYJAGCNYAVMT-VWLOTQADSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: LCMS Assay: The MGAT enzyme reactions were performed in Corning FALCON® 96-well Polypropylene plates, in a total volume of 60 μL of 50 mM Potassium Phosphate buffer pH 7.4, containing a final concentration of 100 μM 2-oleoylglycerol, 15 μM oleoyl-Coenzyme A and 0.0013 μg/μL Human or Mouse MGAT-2 or 0.0026 μg/μL Rat recombinant MGAT-2 membranes expressed in Sf9 cells. Assay plates were run through a fully automated robotics system and shaken for 5 seconds every minute for a total 10 minutes. The reactions were then quenched with 120 μL of ice cold methanol containing 1 μg/mL 1,2-distearoyl-rac-glycerol as the internal standard. Plates were shaken for 2 minutes and spun down to remove protein precipitation. After the spin, samples were transferred to LC/MS compatible PCR plates. For LC/MS analysis, a ThermoFisher Surveyor pump, utilizing a Waters Symmetry C8, 50×2.1 mm column, was used for the chromatography of enzyme products. The buffer system consists of 0.1% formic acid in water with a mobile phase consisting 0.1% formic acid in methanol. The shallow gradient is 90-100% mobile phase in 0.2 min with a total run time of 2.3 min. The first 0.5 minutes of each injection was diverted to waste to eliminate the presence of Phosphate buffer in the enzymatic reaction. The column was run at 0.6 mL/min and a temperature of 65° C.
Measurement: IC50 = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 639.01, ALogP 5.71, HBA 5, HBD 2, PSA 101.57, QED 0.21
Compound (SMILES): O=C(NCCc1c[nH]c2ccc(Cl)cc12)c1cccc(CNC2CCCCC2)c1
InChIKey: TYUBNVSXZSHBQK-UHFFFAOYSA-N
Target: Alpha-synuclein (SINGLE PROTEIN), Homo sapiens
UniProt: P37840
Assay: In Vitro Inhibition Assay: α-Synucleinopathies are characterised by aggregation of α-synuclein in neurons. Aggregation of purified α-synuclein is performed essentially as described by Gerard et al. FASEB. 20(3):524-6 (2006). 20-100 μg purified α-synuclein (Sigma; S7820) at a concentration of about 2.5 μg/mL is incubated in the presence of spermin (250 μM) or paraquat (32 mM) or 6-hydroxydopamine (400 μM) or vehicle in a 384 well plate. Spermin, paraquat and 6-hydroxydopamine promote the α-synuclein aggregation process. Aggregation kinetics is determined by measuring turbidity at 340 nm, every 1-15 minutes for at least one hour.
Measurement: EC50 = 724 nM (pChEMBL 6.139999866485596)
Descriptors: MW 409.96, ALogP 5.22, HBA 2, HBD 3, PSA 56.92, QED 0.5
Compound (SMILES): O=C(N1CCN(c2ccc(N3CCCC3)nn2)CC1)n1sc2c(ccc[n+]2[O-])c1=O
InChIKey: WPYOHTJXPLYYBZ-UHFFFAOYSA-N
Target: Protein-glutamine gamma-glutamyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P21980
Assay: TGM2 Inhibitory Activity Assay: 2 ul of a solution of different concentrations of the test compound in dimethyl sulfoxide was added to 28 ul of buffer (50 mM TRIS, 100 mM NaCl, 10 mM CaCl2, pH 7.5) and 10 ul of a TGM2 solution to give a TMG2 test concentration of 25 ug/ml, and the mixture incubated for 15 minutes at room temperature in a 96 half-well microtiter plate. The enzyme reaction was started by the addition of 10 ul of substrate solution containing H-Abz-APE(CAD-DNP)QEA-OH and glycine methyl ester hydrochloride in buffer to give test concentrations of H-Abz-APE(CAD-DNP)QEA-OH of 50 uM and of glycine methyl ester hydrochloride of 5 mM. The time course of the reaction was monitored with excitation at 318 nm and measurement at emission wavelength 418 nm in a microtiter plate reader (SpectraMax M5, Molecular Devices) over 15 minutes. Rates were calculated from the linear part of the curve (generally between 5 and 10 minutes). IC50 values were calculated from the means (duplicates) of a dilution series of the com
Measurement: IC50 = 140 nM (pChEMBL 6.849999904632568)
Descriptors: MW 427.49, ALogP 0.88, HBA 9, HBD 0, PSA 101.51, QED 0.44
Compound (SMILES): C[C@H](O)CNC(=O)c1cc(NC(=O)c2cc(-c3ncccc3F)c(Cl)cc2Cl)n(-c2ccccc2)n1
InChIKey: ITGHMSHYXORKET-AWEZNQCLSA-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 = 5.78 nM (pChEMBL 8.239999771118164)
Descriptors: MW 528.37, ALogP 4.74, HBA 6, HBD 3, PSA 109.14, QED 0.32
Compound (SMILES): CCNC(=O)Nc1cc(Nc2nccc(C(F)(F)F)n2)cc(-c2ccc([C@@H](O)C(F)F)cc2)c1
InChIKey: GIZANWXTAAKQTO-GOSISDBHSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Homogeneous Time-Resolved Fluorescence (HTRF) Assay: A recombinant GST-hSYK fusion protein was used to measure potency of compounds to inhibit human Syk activity. The recombinant human GST-SYK (Carna Biosciences #08-176) (5 μM final concentration) was incubated with various concentrations of the inhibitor diluted in DMSO (0.1% final concentration) for 10 minutes at room temperature in 15 mM Tris-HCl (pH 7.5), 0.01% tween 20, 2 mM DTT in 384 well plate format. To initiate the reaction the biotinylated substrate peptide (250 nM final concentration) that contains the phosphorylation site for Syk was added with magnesium (5 mM final concentration) and ATP (25 μM final concentration). Final volume of the reaction was 10 μL. Phosphorylation of the peptide was allowed to proceed for 45' at room temperature. To quench the reaction and detect the phosphorylated product, 2 nM of a Europium-anti-phosphotyrosine antibody (Perkin Elmer #AD0161) and 70 nM SA-APC (Perkin-Elmer #CR130-100) were added together in 15 mM Tris pH 7.5, 40 mM EDTA, 0.01% tween 20. Final volume of the quenching solution was 10 μL. The resulting HTRF signal was measured after 30 minutes on an EnVision (Perkin-Elmer) reader using a time-resolved fluorescence protocol.
Measurement: IC50 = 9.9 nM (pChEMBL 8)
Descriptors: MW 481.43, ALogP 5.35, HBA 5, HBD 4, PSA 99.17, QED 0.34
Compound (SMILES): CNC1CCN(C(=O)n2sc3c(ccc[n+]3[O-])c2=O)CC1
InChIKey: SWCBHHMUMMLQKP-UHFFFAOYSA-N
Target: Protein-glutamine gamma-glutamyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P21980
Assay: TGM2 Inhibitory Activity Assay: 2 ul of a solution of different concentrations of the test compound in dimethyl sulfoxide was added to 28 ul of buffer (50 mM TRIS, 100 mM NaCl, 10 mM CaCl2, pH 7.5) and 10 ul of a TGM2 solution to give a TMG2 test concentration of 25 ug/ml, and the mixture incubated for 15 minutes at room temperature in a 96 half-well microtiter plate. The enzyme reaction was started by the addition of 10 ul of substrate solution containing H-Abz-APE(CAD-DNP)QEA-OH and glycine methyl ester hydrochloride in buffer to give test concentrations of H-Abz-APE(CAD-DNP)QEA-OH of 50 uM and of glycine methyl ester hydrochloride of 5 mM. The time course of the reaction was monitored with excitation at 318 nm and measurement at emission wavelength 418 nm in a microtiter plate reader (SpectraMax M5, Molecular Devices) over 15 minutes. Rates were calculated from the linear part of the curve (generally between 5 and 10 minutes). IC50 values were calculated from the means (duplicates) of a dilution series of the com
Measurement: IC50 = 240 nM (pChEMBL 6.619999885559082)
Descriptors: MW 308.36, ALogP 0.35, HBA 6, HBD 1, PSA 81.28, QED 0.61
Compound (SMILES): COCC(=O)NCC(=O)c1c[nH]c2ccc(O)cc12
InChIKey: VZXRQYZONBQNCI-UHFFFAOYSA-N
Target: Sepiapterin reductase (SINGLE PROTEIN), Homo sapiens
UniProt: P35270
Assay: Chromogenic assay: To screen for SPR inhibition, a biochemical assay based on LC/MS (and chromogenic) read-out has been developed. The LC/MS assay monitors the product formation (L-biopterin) and the chromogenic assay measures OD at 420 nm.N-methoxyacetyl serotonin was used as a reference compound (positive control). The IC50 measured using the screening conditions was 20-40 nM, which agrees with the literature (Smith et al., Journal of Biological Chemistry, 297:5601, 1992). The exemplary assay protocol uses the following conditions: SPR (6 nM); L-Sepiapterin (50 uM); NADPH (100 uM); Na-Phosphate buffer, pH 6.5 (100 mM); 82 uL assay volume; 60 minutes incubation with compounds (0.5% final concentration in DMSO) at 37° C. in Greiner uClear 384 well plates.The following experimental procedure was applied: (1) Add 2 uL compound (inhibitor) dilutions (20% DMSO) in Greiner uClear 384 well plates. (2) Add 40 uL enzyme/assay buffer.
Measurement: IC50 = 310 nM (pChEMBL 6.510000228881836)
Descriptors: MW 262.27, ALogP 0.82, HBA 4, HBD 3, PSA 91.42, QED 0.7
Compound (SMILES): COc1cc(-c2cnc(OC)c(Cl)c2)c(F)cc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ncccn3)ccc21
InChIKey: LANRAEKUIXTTLL-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 = 40 nM (pChEMBL 7.400000095367432)
Descriptors: MW 567.99, ALogP 4.45, HBA 9, HBD 1, PSA 125.3, QED 0.31
Compound (SMILES): CCn1cc2ccc(Nc3nc(N[C@@H]4CCCCC[C@@H]4N)nnc3C(N)=O)cc2n1
InChIKey: DIMDKKKMLHGJDY-LSDHHAIUSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Syk Kinase Inhibition Assay: The in vitro inhibitory activity of the compound of the present invention against Syk kinase activity was assayed under the following conditions: the purified human Syk protein used in the test were purchased from Carna Biosciences, Inc. For the inhibitory activity assay on the compound, the compound of the present invention was first serially diluted with dimethyl sulfoxide (DMSO). Next, the purified human Syk protein FL-Peptide 22 (final concentration: 1 uM) (Caliper Life Sciences, Inc.), magnesium chloride (final concentration: 5 mM), ATP (final concentration: 30 uM), and each DMSO solution of the compound of the present invention (final concentration of DMSO: 5%) were added into a buffer solution for reaction (20 mM tris-HCl pH 7.5, 20 mM NaCl, 2.5 mM DTT, 0.02% Tween-20, 1% glycerol, 0.01 mM Pefabloc), and the mixture was then incubated at 25° C. for 30 minutes to perform kinase reaction. The kinase reaction was stopped by the addition thereto of EDTA.
Measurement: IC50 = 0.31 nM (pChEMBL 9.510000228881836)
Descriptors: MW 409.5, ALogP 2.16, HBA 9, HBD 4, PSA 149.66, QED 0.45
Compound (SMILES): COc1cc(OC)cc(N(Cc2nnc(NCCO)o2)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: DTNHNJVOHCLYPS-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 2454.71 nM (pChEMBL 5.610000133514404)
Descriptors: MW 502.54, ALogP 3.17, HBA 12, HBD 2, PSA 136.48, QED 0.29
Compound (SMILES): CC[C@@H](CCC(C)(O)C(F)(F)F)N1C(=O)[C@@](C)(CC(=O)O)C[C@H](c2cccc(Cl)c2)[C@H]1c1ccc(Cl)cc1
InChIKey: GDXYCMHIOJSGMF-ZBBHNODISA-N
Target: E3 ubiquitin-protein ligase Mdm2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00987
Assay: Homogenous Time-Resolved Fluorescence Assay (HTRF1 Assay): The standard assay conditions for the in vitro HTRF assay consisted of a 50 ul total reaction volume in black 384-well Costar polypropylene plates in 1xPBS buffer pH 7.4, 1 mM DTT, 0.1% BSA, 2.5 nM GST-hMDM2 (aa 1-188), 5 nM biotinylated-p53 (aa 1-83), 1.8 nM SA-XLent (Cisbio; Bedford, Mass.), 0.6 nM anti-GST cryptate monoclonal antibody (Cisbio; Bedford, Mass.) and 200 mM KF. Amino acid residues 1-188 of human MDM2 were expressed as an amino-terminal glutathione-S-transferase (GST) fusion protein (GST-hMDM2) in Escherichia coli. Residues 1-83 of human p53 were expressed as an amino-terminal AviTag-TrxA-6-His fusion protein (biotinylated p53) in E. coli. Each protein was purified from cell paste by affinity chromatography.Specifically, 10 uL of GST-hMDM2 was incubated with 10 ul of diluted compound (various concentrations, serially diluted) in 10% DMSO for 20 minutes at room temperature. 20 uL of biotinylated-p53 was added to the GST-hMDM2+compound mixture, and then incubated at room temperature for 60 min. 10 uL of detection buffer consisting of SA-XLent, anti-GST cryptate antibody and KF was added to GST-hMDM2, biotinylated-p53 and compound reaction and left at room temperature to reach equilibrium for >4 hrs. The final concentration of DMSO in the reaction was 2%. Time-resolved fluorescence readings were measured on a microplate multilabel reader. Percentage of inhibition was calculated relative to nutlin-3.
Measurement: IC50 < 10 nM (pChEMBL 8)
Descriptors: MW 574.47, ALogP 7.4, HBA 3, HBD 2, PSA 77.84, QED 0.33
Compound (SMILES): CCNC(=O)c1cc2nccc(Oc3cccc(NC(=O)c4occc4C)c3)c2s1
InChIKey: RLAISKQYVHVVAQ-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Kinase Assay: Kinase assays were performed in 96 well microtiter plates that were coated overnight with 30 μg of poly-Glu-Tyr (4:1) in 10 mM Phosphate Buffered Saline (PBS), pH 7.2-7.4. The plates were incubated with 1% BSA and then washed four times with PBS prior to starting the reaction. Reactions were carried out in 120 μL reaction volumes containing 3.6 μM ATP in kinase buffer (50 mM Hepes buffer pH 7.4, 20 mM MgCl2, 0.1 mM MnCl2 and 0.2 mM Na3VO4). Test compounds were reconstituted in 100% DMSO and added to the reaction to give a final DMSO concentration of 5%. Reactions were initiated by the addition 0.5 ng of purified protein. Following a ten minute incubation at 25 °C., the reactions were washed four times with PBS containing 0.05% Tween-20. 100 μl of a monoclonal anti-phosphotyrosine antibody-peroxidase conjugate was diluted 1:10000 in PBS-Tween-20 and added to the wells for 30 minutes. Following four washes with PBS-Tween-20, 100 μl of 0-phenylenediamine Dihydrochloride in Phosphate-citrate buffer, containing urea hydrogen peroxide, was added to the wells for 7 minutes as a colorimetric substrate for the peroxidase. The reaction was terminated by the addition of 100 μl of 2.5N H2 SO4 to each well and read using a microplate ELISA reader set at 492 nm.
Measurement: IC50 = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 421.48, ALogP 4.99, HBA 6, HBD 2, PSA 93.46, QED 0.46
Compound (SMILES): Cc1nn(C)c(C)c1S(=O)(=O)N1CCC(=C(C#N)c2ccc(Cl)cc2)CC1
InChIKey: YZLYZJPLSNEUDJ-UHFFFAOYSA-N
Target: Prokineticin receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8TCW9
Assay: Antagonist activity at human PKR1 expressed in RBL2H3 cells assessed as inhibition of prokineticin1-mediated intracellular calcium release preincubated for 10 mins followed by PK1 addition by FLIPR assay
Measurement: IC50 = 40 nM (pChEMBL 7.400000095367432)
Descriptors: MW 404.92, ALogP 3.45, HBA 5, HBD 0, PSA 78.99, QED 0.73
Compound (SMILES): CC(C)OCCNC(=O)c1ccc(NC(=O)N2Cc3ccncc3C2)cc1
InChIKey: IPWPYCHMWDRDDA-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 = 20.9 nM (pChEMBL 7.679999828338623)
Descriptors: MW 368.44, ALogP 2.78, HBA 4, HBD 2, PSA 83.56, QED 0.77
Compound (SMILES): CN(C)c1cccc2c(S(=O)(=O)NCc3ccc(NC(=N)c4cccs4)cc3)cccc12
InChIKey: MPMPXVAXSZROEO-UHFFFAOYSA-N
Target: Nitric oxide synthase 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P29476
Assay: Inhibition Assay: The compounds were evaluated for in vitro inhibition against three NOS isozymes: rat nNOS, bovine eNOS and marine iNOS using known literature methods. (Hevel, J. M.; Marietta, M. A. Nitric-oxide synthase assays. Method Enzymol. 1994, 233, 250-258).
Measurement: Ki = 243 nM (pChEMBL 6.610000133514404)
Descriptors: MW 464.62, ALogP 4.88, HBA 5, HBD 3, PSA 85.29, QED 0.27
Compound (SMILES): Cc1ccc(Cl)cc1N1C(=O)c2ccccc2C1(O)c1ccc2nc(Nc3ncccn3)[nH]c2c1
InChIKey: SVFALRKCDRLQHJ-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 482.93, ALogP 4.91, HBA 6, HBD 3, PSA 107.03, QED 0.34
Compound (SMILES): c1cc(-c2n[nH]c3ccc(-c4nnn[nH]4)cc23)ccc1OCCN1CCOCC1
InChIKey: HSZUDLJGFIRCJI-UHFFFAOYSA-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 <= 500 nM (pChEMBL 6.301000118255615)
Descriptors: MW 391.44, ALogP 2.12, HBA 7, HBD 2, PSA 104.84, QED 0.52
Compound (SMILES): O=C(/C=C/c1ccc([N+](=O)[O-])cc1)[n+]1nn([O-])c2ncccc21
InChIKey: VVHQTMAVBQRNPP-VMPITWQZSA-N
Target: Protein-glutamine gamma-glutamyltransferase 2 (SINGLE PROTEIN), Cavia cutleri
UniProt: P08587
Assay: Enzyme Inhibition Assay: For enzymology studies of these compounds, recombinant guinea pig liver TGase was expressed in Escherichia coli and effectively purified (Gillet, S. M. F. G. et al J. N., Prot. Exp. & Purif. 2004, 33, 256). In addition to being easy to obtain in excellent yield and solubility, guinea pig liver TGase was chosen because it shows 80% homology with human tissue TGase (Aeschlimann, D.; Paulsson, M., Throm. Haemost. 1994, 71, 402) and may thus serve as a model for the evaluation of inhibitors of potential therapeutic utility.The IC50 values of synthetic analogues 14a-38a were determined from inhibition of the reaction of 54.4 mM of the chromogenic TGase substrate N-Cbz-Glu(Î³-p-nitrophenyl ester)Gly with 0.010 U of recombinant guinea pig liver TGase as previously reported (Leblanc, A.; Gravel, C.; Labelle, J.; Keillor, J. W. Biochemistry 2001, 40, 8335) and described in detail in the Materials section below. The mode of inhibition was determined for the representative lead compound.
Measurement: IC50 > 20000 nM (pChEMBL 4.698999881744385)
Descriptors: MW 311.26, ALogP 1.33, HBA 7, HBD 0, PSA 117.86, QED 0.31
Compound (SMILES): CNc1nc(Nc2cn(C(C)(C)CO)nc2C)ncc1C(F)(F)F
InChIKey: MRFLTLKAXVFERR-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: Lanthascreen Binding Assay: This assay was used to determine a compound's potency in inhibiting activity of LRRK2 by determining, Kiapp, IC50, or percent inhibition values. In 384 well proxiplates F black, shallow well plates LRRK2, Eu-anti-GST-antibody, Alexa Fluor Kinase tracer 236 and test compound were incubated together.Binding of the Alexa Fluor tracer to a kinase is detected by addition of a Eu-labeled anti-GST antibody. Binding of the tracer and antibody to a kinase results in a high degree of FRET, whereas displacement of the tracer with a kinase inhibitor results in a loss of FRET. Assay conditions and materials used were as follows: Final Assay Conditions: GST-LRRK2 G2019S 10 nM Eu-anti-GST-antibody 2 nM Kinase tracer 236 8.5 nM Kinase reaction time: 1 hour Temperature: ambient Total volume: 15 ul DMSO 1% Materials: 384 well proxiplates F black shallow cat# 6008260 well Perkin Elmer Kinase: LRRK2 G2019S, Invitrogen cat # PV4882(LOT 567054A).
Measurement: Ki = 4.8 nM (pChEMBL 8.319999694824219)
Descriptors: MW 344.34, ALogP 2.51, HBA 7, HBD 3, PSA 87.89, QED 0.77
Compound (SMILES): Cc1cccc(C)c1N1C[C@@](C(=O)Nc2cc(C(F)(F)F)cc(C(F)(F)F)c2)(C(C)C)[C@H](C)C1=O
InChIKey: MCGYLBXVWYMXIB-WAIKUNEKSA-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 500.48, ALogP 6.6, HBA 2, HBD 1, PSA 49.41, QED 0.48
Compound (SMILES): O=C(Nc1ccc(CNc2nc(N3CCCC3)nc3ccc(C(F)(F)F)cc23)cc1)c1ccc(Cl)cc1
InChIKey: KBTMSHBWOZXTIL-UHFFFAOYSA-N
Target: TCF4/beta-catenin (PROTEIN COMPLEX), Homo sapiens
UniProt: P35222
Assay: Inhibition of TCF-4/beta catenin in human Tcf33.13 cells expressing Tcf-4/SV40 binding sites driving firefly luciferase reporter gene after 24 hrs by luminometer
Measurement: IC50 = 6554 nM (pChEMBL 5.179999828338623)
Descriptors: MW 525.96, ALogP 6.77, HBA 5, HBD 2, PSA 70.15, QED 0.29
Compound (SMILES): COc1ccc([C@@H](CO)NC(=O)c2ccc(NC(=O)N3Cc4ccccc4C3)cc2)cc1
InChIKey: NTHLOULJQMNXGT-HSZRJFAPSA-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 = 21.4 nM (pChEMBL 7.670000076293945)
Descriptors: MW 431.49, ALogP 3.71, HBA 4, HBD 3, PSA 90.9, QED 0.56
Compound (SMILES): Cc1c(C(=O)Nc2c(C)n(C)n(C3CCCCC3)c2=O)noc1C1CCC2(CC1)OCCO2
InChIKey: CWAHGMBCLDRHIJ-UHFFFAOYSA-N
Target: E3 ubiquitin-protein ligase SMURF1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9HCE7
Assay: Inhibition of SMURF1 (unknown origin) in presence of E1/E2/Ub mix measured after 45 mins by europium-labeled streptavidin based TR-FRET assay
Measurement: IC50 = 420 nM (pChEMBL 6.380000114440918)
Descriptors: MW 458.56, ALogP 3.95, HBA 8, HBD 1, PSA 100.52, QED 0.74
Compound (SMILES): CC(O)c1cccc(C#Cc2c(N)ncnc2N[C@@H](C)c2nc3cccc(Cl)c3c(=O)n2-c2ccccc2)n1
InChIKey: DILPCNSPSXCYPR-ZENAZSQFSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42336
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Assay: PI3K isoforms were assayed under initial rate conditions in the presence of 25 mM Hepes (pH 7.4), and 2xKm ATP (100-300 uM), 10 uM PIP2, 5% glycerol, 5 mM MgCl2, 50 mM NaCl, 0.05% (v/v) Chaps, 1 mM dithiothreitol, 1% (v/v) DMSO at the following concentrations for each isoform: PI3K alpha, beta, delta, and gamma at 50 picomolar (pM) and PI3Kgamma at 2 nanomolar (nM). After an assay reaction time of 30 minutes at 25° C., reactions were terminated with a final concentration of 10 mM EDTA, 10 nM labeled-PIP3, and 35 nM Europium labeled GRP-1 detector protein before reading TR-FRET on an Envision plate reader (Ex: 340 nm; Em: 615/665 nm; 100 us delay and 500 us read window).
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 538.01, ALogP 4.43, HBA 9, HBD 3, PSA 131.84, QED 0.28
Compound (SMILES): CC(C)Nc1cc(-n2ncc3cc(C#N)cnc32)ncc1C(=O)NC1CCN(C(=O)C(C)C)CC1
InChIKey: DLYNLVXDPMFCGF-UHFFFAOYSA-N
Target: Interleukin-1 receptor-associated kinase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NWZ3
Assay: Inhibition Assay: The assays were performed in U-bottom 384-well plates. The final assay volume was 30 L prepared from 15 L additions of enzyme and substrates (fluoresceinated peptide and ATP) and test compounds in assay buffer (20 mM HEPES pH 7.2, 10 mM MgCl2, 0.015% Brij 35 and 4 mM DTT). The reaction was initiated by the combination of IRAK4 with substrates and test compounds. The reaction mixture was incubated at room temperature for 60 min. and terminated by adding 45 μL of 35 mM EDTA to each sample. The reaction mixture was analyzed on the Caliper LABCHIP® 3000 (Caliper, Hopkinton, Mass.) by electrophoretic separation of the fluorescent substrate and phosphorylated product. Inhibition data were calculated by comparison to no enzyme control reactions for 100% inhibition and vehicle-only reactions for 0% inhibition. The final concentrations of reagents in the assays are ATP, 500 μM; FL-IPTSPITTTYFFFKKK peptide 1.5 μM; IRAK4, 0.6 nM; and DMSO, 1.6%.
Measurement: IC50 = 18.6 nM (pChEMBL 7.730000019073486)
Descriptors: MW 474.57, ALogP 2.88, HBA 8, HBD 2, PSA 128.83, QED 0.56
Compound (SMILES): COc1cc(C(=O)Nc2cnn(Cc3c(C)noc3C)c2)cc(O)c1O
InChIKey: UPUBFWVIQLTVKL-UHFFFAOYSA-N
Target: Taste receptor type 2 member 8 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NYW2
Assay: Fluorescence Polarization Assays: In another embodiment, Fluorescence Polarization ("FP") based assays may be used to detect and monitor ligand binding. Fluorescence polarization is a versatile laboratory technique for measuring equilibrium binding, nucleic acid hybridization, and enzymatic activity. Fluorescence polarization assays are homogeneous in that they do not require a separation step such as centrifugation, filtration, chromatography, precipitation, or electrophoresis. These assays are done in real time, directly in solution and do not require an immobilized phase. Polarization values can be measured repeatedly and after the addition of reagents since measuring the polarization is rapid and does not destroy the sample. Generally, this technique can be used to measure polarization values of fluorophores from low picomolar to micromolar levels.
Measurement: IC50 = 2200 nM (pChEMBL 5.659999847412109)
Descriptors: MW 358.35, ALogP 2.21, HBA 8, HBD 3, PSA 122.64, QED 0.6
Compound (SMILES): CCOc1cc2ncnc(Nc3ccc(OCc4ccccn4)cc3)c2cc1NC(=O)/C=C/CN1CCCC1
InChIKey: CZRDEUGQKBEMMH-VQHVLOKHSA-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 = 98 nM (pChEMBL 7.010000228881836)
Descriptors: MW 524.63, ALogP 5.34, HBA 8, HBD 2, PSA 101.5, QED 0.25
Compound (SMILES): COc1cccc2c1ncc1c(=O)n(C)c(=O)n(C3CCCC3)c12
InChIKey: AIQDEWPLFZXAOT-UHFFFAOYSA-N
Target: Tumor necrosis factor (SINGLE PROTEIN), Homo sapiens
UniProt: P01375
Assay: ELISA Assay: Add 100 μL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature. Repeat the aspiration/wash as in step 2 of Plate Preparation. Add 100 μL of the Detection Antibody, diluted in Reagent Diluent, to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature. Repeat the aspiration/wash as in step 2 of Plate Preparation. Add 100 μL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light. Repeat the aspiration/wash as in step 2. Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light. Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing. Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.
Measurement: IC50 = 1400 nM (pChEMBL 5.849999904632568)
Descriptors: MW 325.37, ALogP 2.37, HBA 6, HBD 0, PSA 66.12, QED 0.68
Compound (SMILES): NS(=O)(=O)c1ccc(O[C@H]2c3ccccc3C[C@@H]2N2CCCC2)c(Cl)c1Cl
InChIKey: WGVDATSCZYCIBM-LIRRHRJNSA-N
Target: Cytochrome P450 2D6 (SINGLE PROTEIN), Homo sapiens
UniProt: P10635
Assay: Inhibition of CYP2D6 in pooled human hepatic microsomes using dextromethorphan substrate in presence of NADPH
Measurement: IC50 = 400 nM (pChEMBL 6.400000095367432)
Descriptors: MW 427.35, ALogP 3.78, HBA 4, HBD 1, PSA 72.63, QED 0.81
Compound (SMILES): Nc1ncc(-c2ccc3ccccc3c2)c(-c2ccccc2)n1
InChIKey: OLHJFBKEAWWWSM-UHFFFAOYSA-N
Target: Adenosine receptor A2a (SINGLE PROTEIN), Homo sapiens
UniProt: P29274
Assay: Displacement of [3H]ZM241385 from recombinant human adenosine A2A receptor expressed in HEK293 cell membranes after 2 hrs by beta scintillation counting method
Measurement: Ki = 129 nM (pChEMBL 6.889999866485596)
Descriptors: MW 297.36, ALogP 4.55, HBA 3, HBD 1, PSA 51.8, QED 0.59
Compound (SMILES): O=C(Nc1ccc(C2=CCN(C(=O)C3COCCO3)CC2)cc1)N1Cc2ccncc2C1
InChIKey: LSXSNCQDNSNHPF-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 = 5.73 nM (pChEMBL 8.239999771118164)
Descriptors: MW 434.5, ALogP 2.66, HBA 5, HBD 1, PSA 84, QED 0.8
Compound (SMILES): COc1cc2c(Oc3ccc(NC(=O)c4cn(-c5ccccc5OC(F)(F)F)cn4)cc3F)ccnc2cc1OCCCN1CCN(C)CC1
InChIKey: GHKBAXBRMKMFEB-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Inhibition of c-MET (unknown origin) using poly (Glu, Tyr) 4:1 as substrate after 30 mins by HTRF assay
Measurement: IC50 = 43 nM (pChEMBL 7.369999885559082)
Descriptors: MW 694.69, ALogP 6.53, HBA 10, HBD 1, PSA 103.21, QED 0.12
Compound (SMILES): C[C@H](O)[C@@H](O)c1cnc(N2CCN(C(=O)Nc3nc4cc(F)ccc4s3)[C@@H](C)C2)c(F)c1
InChIKey: KJXOKUPEUUCKKK-OAVHHTNSSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8NER1
Assay: Capsaicin-based Assay: One day prior to assay, TRPV1/CHO cells were seeded in 96-well clear-bottom black plates (20,000 cells/well) in growth media. On the day of the experiment, the cells were washed with 0.2 mL 1x Hank's Balanced Salt Solution (Life Technologies) containing 1.6 mM CaCl2 and 20 mM HEPES, pH 7.4 ("wash buffer"). Subsequently, the cells were loaded by incubation in 0.1 mL of wash buffer containing Fluo-4 at 3 uM final concentration. After 1 hour, the cells were washed twice with 0.2 mL wash buffer and resuspended in 0.1 mL wash buffer. The plates were then transferred to a Fluorescence Imaging Plate Reader (Molecular Devices). Fluorescence intensity was monitored for 15 seconds to establish a baseline. Subsequently, test compounds diluted in assay buffer (1x Hank's Balanced Salt Solution containing 1 mM CaCl2 and 20 mM HEPES, pH 7.4) containing 1% DMSO were added to the cell plate and fluorescence was monitored for 2 minutes. The final concentration of the compound was adjusted to range from 100 uM to 1.5625 uM. If the test compound was an especially potent antagonist, the final concentration of the compound was adjusted to range from 10 uM to 1.5625 nM. Human TRPV1 was then activated by the addition of 50 uL capsaicin (100 nM final concentration) and plates incubated for an additional 3 min.
Measurement: IC50 = 107 nM (pChEMBL 6.96999979019165)
Descriptors: MW 463.51, ALogP 3.13, HBA 7, HBD 3, PSA 101.82, QED 0.55
Compound (SMILES): NC[C@H]1CC[C@H](C(=O)N2CCc3ccccc3[C@H]2C(=O)Nc2ccc(C(=O)O)cc2)CC1
InChIKey: VVENGVCWAPTCEF-ZJBJCVSYSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: In Vitro Assay: Factor XIa determinations were made in 50 mM HEPES buffer at pH 7.4 containing 145 mM NaCl, 5 mM KCl, and 0.1% PEG 8000 (polyethylene glycol; JT Baker or Fisher Scientific). Determinations were made using purified human Factor XIa at a final concentration of 75-200 pM (Haematologic Technologies) and the synthetic substrate S-2366 (pyroGlu-Pro-Arg-pNA; CHROMOGENIX® or AnaSpec) at a concentration of 0.0002-0.001 M.
Measurement: Ki = 84.61 nM (pChEMBL 7.070000171661377)
Descriptors: MW 435.52, ALogP 3.21, HBA 4, HBD 3, PSA 112.73, QED 0.67
Compound (SMILES): Cc1ccc(NC(=O)c2cccc(C(F)(F)F)c2)cc1-c1cc(N2CCOCC2)nc(NS(C)(=O)=O)n1
InChIKey: CWPVMCLRIZDTFY-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 = 3.454 nM (pChEMBL 8.460000038146973)
Descriptors: MW 535.55, ALogP 3.93, HBA 7, HBD 2, PSA 113.52, QED 0.49
Compound (SMILES): C[C@@H](CCCCc1ccccc1)[C@@H](O)/C=C/[C@H]1CC(F)(F)C(=O)N1CCCc1ccc(C(=O)O)s1
InChIKey: WQNDXGHRPMQDFT-BQMVMUGMSA-N
Target: Prostaglandin E2 receptor EP4 subtype (SINGLE PROTEIN), Homo sapiens
UniProt: P35408
Assay: cAMP Assay: 1. Seed cells on an EP2 or EP4 STEP plate at a density of 40,000-80,000 cells/well in 200 uL of reduced serum medium containing 0.5% FBS. Place the plate in a 37° C. incubator with 5% CO2 and incubate overnight. 2. After 16-18 hours of incubation, aspirate the culture media from each well. 3. Add 200 ul of culture medium containing 500 uM IBMX (an inhibitor of cAMP phosphodiesterase) and different concentration of test compounds to the assigned wells. For each test compound, at least 8 concentrations starting at highest 10 M and lowest 0.01 pM were tested. In addition each concentration had triplicates. A PGE2 curve (concentrations from lowest to highest, 0 pM, 0.384 pM, 1.92 pM, 9.6 pM, 48 pM, 240 pM, 1200 pM, and 6000 pM) was always run in parallel with test compounds. 4. Incubate the cells in a cell culture incubator for 30 minutes. 5. Centrifuge the plate at 1,000x rpm for 10 minutes. 6. Aspirate the supernatant.
Measurement: EC50 = 0.37 nM (pChEMBL 9.430000305175781)
Descriptors: MW 505.63, ALogP 5.58, HBA 4, HBD 2, PSA 77.84, QED 0.28
Compound (SMILES): CN1C(=O)N(c2cccc(C(F)(F)F)c2)C2=C(C(=O)CC2)C1c1ccc(C#N)cc1C(=O)NCc1cn[nH]c1
InChIKey: JGKRILWFOJPJCY-UHFFFAOYSA-N
Target: Neutrophil elastase (SINGLE PROTEIN), Homo sapiens
UniProt: P08246
Assay: Human Neutrophil Elastase Assay: The following buffers were used: 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.Assay conditions: 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 μL 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 = 5 nM (pChEMBL 8.300000190734863)
Descriptors: MW 534.5, ALogP 4.46, HBA 5, HBD 2, PSA 122.19, QED 0.5
Compound (SMILES): O=C(Nc1nc2ccccc2s1)c1cccc2c1CN(c1ccc(-c3cnn(Cc4cccnc4)c3)c(C(=O)O)n1)CC2
InChIKey: KFWWHDXQQKVMHS-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q07817
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The measurement of competition of compounds of Formula (I) with F-Bak for a Bcl-2 family protein (Bcl-xL) binding site using a Time Resolved Fluorescence Resonance Energy Transfer (TR-FRET) binding assay: Test compounds were serially diluted in DMSO starting at 50 M (2x starting concentration; 10% DMSO) and 10 Î¼L transferred into a 384-well plate. Then 10 Î¼L of a protein/probe/antibody mix is added to each well at final concentrations listed in Table 1. Protein: GST-Bcl-xL; 1(nM) Probe: F-Bak (GQVGRQLAIIGDK (6-FAM)INR-amide) SEQ ID NO: 1; 100(nM) Antibody: Tb-anti-GST; 1(nM). The samples are then mixed on a shaker for 1 minute then incubated for an additional 2 hours at room temperature. For each assay plate, a probe/antibody and protein/antibody/probe mixture were included as a negative and a positive control, respectively. Fluorescence was measured on the ENVISION plate reader (Perkin Elmer) using a 340/35 nm excitation filter and 520/525 (F-Bak) and 495/510 nm (Tb-labeled anti-his antibody) emission filters. Dissociation constants (Ki) were determined using Wang's equation (Wang, Z.X. An 20 exact mathematical expression for describing competitive binding of two different ligands to protein molecule. FEBS Lett. 1995 360:111-114). The TR-FRET assay can be performed in the presence of varying concentrations of human serum (HS) or fetal bovine serum (FBS).
Measurement: Ki = 258 nM (pChEMBL 6.590000152587891)
Descriptors: MW 587.67, ALogP 5.51, HBA 9, HBD 2, PSA 126.13, QED 0.25
Compound (SMILES): Cc1cc(Nc2nccc(C(F)(F)F)n2)cc(-c2cnc(C(F)C3CCC(C(=O)O)CC3)s2)c1
InChIKey: OHQHGRYHFAMFJU-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Homogeneous Time-Resolved Fluorescence (HTRF) Assay: A recombinant GST-hSyk fusion protein was used to measure potency of compounds to inhibit human Syk activity. The recombinant human GST-Syk (Carna Biosciences #08-176) (5 pM final concentration) was incubated with various concentrations of the inhibitor diluted in DMSO (0.1% final concentration) for 10 minutes at room temperature in 15 mM Tris-HCl (pH 7.5), 0.01% tween 20, 2 mM DTT in 384 well plate format. To initiate the reaction the biotinylated substrate peptide (250 nM final concentration) that contains the phosphorylation site for Syk was added with magnesium (5 mM final concentration) and ATP (25 μM final concentration). Final volume of the reaction was 10 μL. Phosphorylation of the peptide was allowed to proceed for 45′ at room temperature. To quench the reaction and detect the phosphorylated product, 2 nM of a Europium-anti-phosphotyrosine antibody (Perkin Elmer #AD0161) and 70 nM SA-APC (Perkin-Elmer #CR130-100) were added together in 15 mM Tris pH 7.5, 40 mM
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 494.51, ALogP 6.57, HBA 6, HBD 2, PSA 88, QED 0.38
Compound (SMILES): CC(O)c1ccc2[nH]c(-c3nc(C(F)(F)F)sc3N3CCN(C(=O)Cn4cnc5cccnc54)[C@H](C)C3)nc2c1
InChIKey: GUACISYFMJNFES-GICMACPYSA-N
Target: C-X-C chemokine receptor type 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P49682
Assay: FLIPR Assay: The bioactivity of compounds is tested in a fluorometric imaging plate reader (FLIPR: Molecular Devices) using engineered CHO-K1 cells expressing the human CXCR3A coupled to a G protein (Galpha(16)). Cells are plated the day prior to bioassay in F12 medium supplemented with 10% FBS and G418 and hygromycin antibiotics to maintain recombinant selection. At the day of bioassay, cells are washed and dye loaded for one hour with Fluo-4-AM (Invitrogen) in Hanks Balanced Salt Solution (Invitrogen), buffered with 20 mM Hepes at pH 7.4 and sodium bicarbonate (0.015%), containing 5 mM probenecid. This buffer, but lacking the dye and containing probenecid at a concentration of 2.5 nM, is also is used for washing steps (wash buffer); or lacking both dye and probenecid but supplemented with 0.1% BSA for compound dilution steps (dilution buffer). Cells are washed free of excess dye and 60 microliter of wash buffer is added. Stock solutions of test compounds are made up at a concentration of 10 mM in DMSO, and serially diluted in dilution buffer to concentrations required for inhibition dose response curves. After a 10 minute incubation period at 37° C., 10 microliters of each compound dilution are transferred from a compound plate to the plate containing the recombinant cells in the FLIPR instrument according to the manufacturer's instructions. Following basal readings, 10 microliter CXCL10 agonist at a concentration of 20 nM (from Peprotech) is added, again using the FLIPR instrument. Changes in fluorescence are monitored before and after addition of the test compounds.
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 570.6, ALogP 4.24, HBA 9, HBD 2, PSA 116.06, QED 0.32
Compound (SMILES): CCC(CNC(=O)c1ccc(NC(=O)N2Cc3ccccc3C2)cc1)OC
InChIKey: VZDKIYYJBVBNJD-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 = 11.6 nM (pChEMBL 7.940000057220459)
Descriptors: MW 367.45, ALogP 3.39, HBA 3, HBD 2, PSA 70.67, QED 0.82
Compound (SMILES): CCOc1cc(C(=O)N2CC(OC)C2)ccc1Nc1cc2cc(-c3cnn(CC(C)(C)O)c3)ccc2cn1
InChIKey: KDQSBJSLGKAMSI-UHFFFAOYSA-N
Target: Dual specificity protein kinase TTK (SINGLE PROTEIN), Homo sapiens
UniProt: P33981
Assay: Kinase Assay: The enzyme reaction (total volume 10 μl) was carried out in black 384-well low volume plates containing full length MPS1 (12.5 nM or 3 nM), fluorescent labelled peptide [known as H236, which has the sequence: 5FAM-DHTGFLTEYVATR-CONH2](5 μM), ATP (10 μM), either DMSO (1% v/v) or the test compound (in the range 0.25 nM-100 μM in 1% DMSO) and assay buffer (50 mM HEPES (pH 7.0), 0.02% NaN3, 0.01% BSA, 0.1 mM Orthovandate, 10 μM MgCl2, 1 μM DTT, Roche protease inhibitor). The reaction was carried out for 60 min at room temperature and stopped by the addition of buffer (10 μl) containing 20 mM EDTA, 0.05% (v/v) Brij-35, in 0.1M HEPES-buffered saline (Free acid, Sigma, UK). The plate was read on a Caliper EZ reader II (Caliper Life Sciences).
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 515.61, ALogP 4.48, HBA 8, HBD 2, PSA 101.74, QED 0.34