lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CN[C@@H](C)C(=O)N[C@H]1COc2ccccc2N(Cc2c(Cl)n(Cc3ccccc3)c3ccccc23)C1=O
InChIKey: IOHRCWWNKMUJHT-CVDCTZTESA-N
Target: E3 ubiquitin-protein ligase XIAP (SINGLE PROTEIN), Homo sapiens
UniProt: P98170
Assay: TR-FRET Assay: Ten nanomolar of 6x Histidine-tagged BIR2 domain, corresponding to amino acids 124-240 of XIAP, or BIR3 domain, corresponding to amino acids 241-356 of XIAP, was mixed with 20 nM of the peptide AVPIAQKSEK-(8-biotin)-OH 1:2 TFA, in the presence of 50 mM Tris-C1, pH 7.5, 100 mM NaCl, 1 mM dithiothreitol (DTT) and 0.1 mg/mL bovine serum albumin (BSA). Following a 45 min. incubation at 37° C., Europium-Streptavidin and Allophycocyanin conjugated anti-Histidine antibody were added to a final concentration of 1.5 nM and 15 nM, respectively. Time-resolved fluorescence resonance energy transfer (TR-FRET) signals were measured 1 hour later at room temperature. Test compound potency was assessed at 10 serially diluted concentrations.
Measurement: IC50 = 167 nM (pChEMBL 6.78000020980835)
Descriptors: MW 517.03, ALogP 4.36, HBA 5, HBD 2, PSA 75.6, QED 0.38
Compound (SMILES): CC#CCOc1ccc(C(=O)Nc2ccc(F)c([C@]3(C)CS(=O)(=O)C4(CN(C)C4)C(=N)N3)c2)nc1
InChIKey: QLVXMZONSZVPIM-QHCPKHFHSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: HTRF FRET Assay: A homogeneous time-resolved FRET assay can be used to determine IC50 values for inhibitors of the soluble human BACE1 catalytic domain. This assay monitors the increase of 620 nm fluorescence that resulted from BACE1 cleavage of an APPswedish APPswe mutant peptide FRET substrate (QSY7-EISEVNLDAEFC-Europium-amide). This substrate contains an N-terminal QSY7 moiety that serves as a quencher of the C-terminal Europium fluorophore (620 nm Em). In the absence of enzyme activity, 620 nm fluorescence is low in the assay and increased linearly over 3 hours in the presence of uninhibited BACE1 enzyme. Inhibition of BACE1 cleavage of the QSY7-APPswe-Eu substrate by inhibitors is manifested as a suppression of 620 nm fluorescence.Varying concentrations of inhibitors at 3x the final desired concentration in a volume of 10 ul are preincubated with purified human BACE1 catalytic domain (3 nM in 10 ul) for 30 minutes at 30° C. in reaction buffer containing 20 mM Na-Acetate pH 5.0, 10% glycerol, 0.1% Brij-35 and 7.5% DSMO. Reactions are initiated by addition of 10 ul of 600 nM QSY7-APPswe-Eu substrate (200 nM final) to give a final reaction volume of 30 ul in a 384 well Nunc HTRF plate. The reactions are incubated at 30° C. for 1.5 hours. The 620 nm fluorescence is then read on a Rubystar HTRF plate reader (BMG Labtechnologies) using a 50 millisecond delay followed by a 400 millisecond acquisition time window. Inhibitor IC50 values are derived from non-linear regression analysis of concentration response curves. Ki values are then calculated from IC50 values using the Cheng-Prusoff equation using a previously determined um value of 8 uM for the QSY7-APPswe-Eu substrate at BACE1.
Measurement: Ki = 2.3 nM (pChEMBL 8.640000343322754)
Descriptors: MW 499.57, ALogP 1.77, HBA 7, HBD 3, PSA 124.48, QED 0.54
Compound (SMILES): CCS(=O)(=O)Nc1n[nH]c2ccc(-c3cn(Cc4ccccc4)nn3)cc12
InChIKey: LLGGBSASIYAINF-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 = 410 nM (pChEMBL 6.389999866485596)
Descriptors: MW 382.45, ALogP 2.63, HBA 6, HBD 2, PSA 105.56, QED 0.53
Compound (SMILES): COc1cc(-c2ccc(Cl)cc2F)c(F)cc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: GMXZBWWMPJQNCZ-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 = 20 nM (pChEMBL 7.699999809265137)
Descriptors: MW 543.94, ALogP 5.39, HBA 7, HBD 1, PSA 103.43, QED 0.31
Compound (SMILES): O=c1c2cn[nH]c2c2ccc(-c3cccc(N4CCOCC4)n3)cc2n1CC(F)(F)F
InChIKey: ISSPVKARXSMUCN-UHFFFAOYSA-N
Target: Methionine aminopeptidase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P50579
Assay: Enzymatic Screening Assay: The enzymatic test is a test in two steps.In a first step, it consists in placing in contact the compound according to the invention, the dialysed MetAP2 protein and the substrate (Met-Pro-Arg-pNa peptide synthesized by Neosystem), the N-terminal methionine of which can be cleaved with MetAP2, and which bears at the C-terminal end a para-nitroaniline (pNa) chromophore, which can itself be released by another peptidase only when the N-terminal methionine has been cleaved beforehand.Consequently, the second step consists in reacting the peptides cleaved in the preceding step with a second peptidase in order to release the chromophore. The peptidase used in this second step is cathepsin, which comes from the TagZyme DAPase kit (Quiagen, 34366).The MetAP2 activity is proportional to the amount of para-nitroaniline released, which is measured by absorbance at 405 nm.
Measurement: IC50 = 362 nM (pChEMBL 6.440000057220459)
Descriptors: MW 429.4, ALogP 3.34, HBA 6, HBD 1, PSA 76.04, QED 0.54
Compound (SMILES): Cc1cc(F)cc(C)c1N1CC(C(=O)Nc2cc(C(F)(F)F)cc(C(F)(F)F)c2)CC1=O
InChIKey: MKYZYIZITJDHFT-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 462.37, ALogP 5.47, HBA 2, HBD 1, PSA 49.41, QED 0.62
Compound (SMILES): O=C(Nc1cccc(Cc2n[nH]c(=O)c3c2CCCC3)c1)C1CCN1
InChIKey: UVUKTIGEZULZMN-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P09874
Assay: PARP Enzyme Inhibition Assay: PARP1 assay was conducted in PARP assay buffer containing 50 mM Tris pH 8.0, 1 mM DTT, 4 mM MgCl2. PARP reactions contained 1.5 uM [3H]-NAD+ (1.6 uCi/mmol), 200 nM biotinylated histone H1, 200 nM sIDNA, and 1 nM PARP enzyme. Auto reactions utilizing SPA bead-based detection were carried out in 100 uL volumes in white 96 well plates. Reactions were initiated by adding 50 ul of 2xNAD+ substrate mixture to 50 uL of 2x enzyme mixture containing PARP and DNA. These reactions were terminated by the addition of 150 uL of 1.5 mM benzamide (~1000-fold over its IC50). 170 uL of the stopped reaction mixtures were transferred to streptavidin Flash Plates, incubated for 1 hour, and counted using a TopCount microplate scintillation counter.
Measurement: Ki = 136 nM (pChEMBL 6.869999885559082)
Descriptors: MW 338.41, ALogP 1.54, HBA 4, HBD 3, PSA 86.88, QED 0.79
Compound (SMILES): COc1ccccc1Cn1cc(CCCCC[C@@H]2SC[C@@H]3NC(=O)N[C@@H]32)nn1
InChIKey: WIVRTUIOWOVNAH-WDSOQIARSA-N
Target: Biotin--protein ligase (SINGLE PROTEIN), Homo sapiens
UniProt: P50747
Assay: Inhibition of human biotin protein ligase assessed as reduction in 3H-biotin incorporation into pyruvate carboxylase biotin domain after 10 mins by liquid scintillation counting analysis
Measurement: Ki > 16000 nM (pChEMBL 4.796000003814697)
Descriptors: MW 401.54, ALogP 2.6, HBA 6, HBD 2, PSA 81.07, QED 0.5
Compound (SMILES): CC1CCN(C(=O)C(CCn2cccc2C#N)NS(=O)(=O)c2ccc3cc[nH]c3c2)CC1
InChIKey: AFQNQMQRHXUWTJ-UHFFFAOYSA-N
Target: C-C chemokine receptor type 10 (SINGLE PROTEIN), Homo sapiens
UniProt: P46092
Assay: Antagonist activity at human CCR10 expressed in CHOK1 cells coexpressing aequorin/Galphaq assessed as inhibition of human CCL27-dependent calcium flux in presence of coelenterazine H by chemiluminescence assay
Measurement: IC50 = 7.943 nM (pChEMBL 8.100000381469727)
Descriptors: MW 453.57, ALogP 2.84, HBA 5, HBD 2, PSA 110.99, QED 0.57
Compound (SMILES): O=C(Nc1sc(-c2ccc(CN3CC(F)C3)cc2)nc1-c1cc(Cl)ccc1OC(F)F)c1cnn2cccnc12
InChIKey: QJPQQWJPPVTUDB-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: JAK Enzyme Assay: The activity of the isolated recombinant JAK1 and JAK2 kinase domain was measured by monitoring phosphorylation of a peptide derived from JAK3 (Val-Ala-Leu-Val-Asp-Gly-Tyr-Phe-Arg-Leu-Thr-Thr, fluorescently labeled on the N-terminus with 5-carboxyfluorescein) using the Caliper LabChip® technology (Caliper Life Sciences, Hopkinton, Mass.). To determine inhibition constants (Ki), compounds were diluted serially in DMSO and added to 50 μL kinase reactions containing purified enzyme (1.5 nM JAK1, or 0.2 nM JAK2), 100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 1.5 μM peptide substrate, ATP (25 μM), 10 mM MgCl2, 4 mM DTT at a final DMSO concentration of 2%. Reactions were incubated at 22° C. in 384-well polypropylene microtiter plates for 30 minutes and then stopped by addition of 25 μL of an EDTA containing solution (100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 150 mM EDTA), resulting in a final EDTA concentration of 50 mM. After termination of the kinase reaction, the proportion of phosphorylated product was determined as a fraction of total peptide substrate using the Caliper LabChip® 3000 according to the manufacturer's specifications.
Measurement: Ki = 1.31 nM (pChEMBL 8.880000114440918)
Descriptors: MW 585.01, ALogP 6.18, HBA 8, HBD 1, PSA 84.65, QED 0.23
Compound (SMILES): COCCC1(Cn2ncc(-c3ccc(N4CCc5cccc(C(=O)Nc6nc7ccccc7s6)c5C4)nc3C(=O)O)c2C)CCCCCC1
InChIKey: DGPHUOWUXMACOK-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 < 0.1 nM (pChEMBL 10)
Descriptors: MW 678.86, ALogP 7.75, HBA 9, HBD 2, PSA 122.47, QED 0.14
Compound (SMILES): COc1ccccc1CN(Cc1ccco1)S(=O)(=O)c1ccc(C(=O)O)cc1
InChIKey: UOCGGLUFMCGMCT-UHFFFAOYSA-N
Target: Taste receptor type 2 member 14 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NYV8
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 = 12000 nM (pChEMBL 4.920000076293945)
Descriptors: MW 401.44, ALogP 3.38, HBA 5, HBD 1, PSA 97.05, QED 0.62
Compound (SMILES): CNC(=O)c1c(-c2ccc(F)cc2)oc2cc(N(C)S(C)(=O)=O)c(-c3ccc4c(n3)-c3nn5ccccc5c3CO4)cc12
InChIKey: VPKMRTUSLGYLDC-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1a (isolate H) (HCV)
UniProt: P27958
Assay: ELISA-Based Assay: Measurement of inhibition by compounds was performed using the HCV replicon system. Several different replicons encoding different HCV genotypes or mutations were used. In addition, potency measurements were made using different formats of the replicon assay, including different ways of measurements and different plating formats. See Jan M. Vrolijk et al., A replicons-based bioassay for the measurement of interferons in patients with chronic hepatitis C, 110 J. VIROLOGICAL METHODS 201 (2003); Steven S. Carroll et al., Inhibition of Hepatitis C Virus RNA Replication by 2'-Modified Nucleoside Analogs, 278(14) J. BIOLOGICAL CHEMISTRY 11979 (2003). However, the underlying principles are common to all of these determinations, and are outlined below.Stable neomycin phosphotransferase encoding replicons-harboring cell lines were used, so all cell lines were maintained under G418 selection prior to the assay. Potency was determined using a cell ELISA assay with an antibody to the replicons encoded NS3/4a protease. See Caterina Trozzi et al., In Vitro Selection and Characterization of Hepatitis C Virus Serine Protease Variants Resistant to an Active-Site Peptide Inhibitor, 77(6) J. Virol. 3669 (2003).
Measurement: IC50 = 225 nM (pChEMBL 6.650000095367432)
Descriptors: MW 597.63, ALogP 5.26, HBA 8, HBD 1, PSA 119.04, QED 0.29
Compound (SMILES): CNC(=O)c1cccc(Nc2nccc(N3CC(Oc4ccc(Cl)cc4)C3)n2)c1
InChIKey: SJYSXEUCXOFRPG-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 = 430 nM (pChEMBL 6.369999885559082)
Descriptors: MW 409.88, ALogP 3.5, HBA 6, HBD 2, PSA 79.38, QED 0.65
Compound (SMILES): CCOc1ccc(C[C@@H](NC(=O)c2ccccc2)C(=O)N[C@@H](Cc2cccc(F)c2)C(=O)NCc2ccc(CN)cc2)cc1
InChIKey: OYMHEDPIXZJWGT-AJQTZOPKSA-N
Target: Plasma kallikrein (SINGLE PROTEIN), Homo sapiens
UniProt: P03952
Assay: In Vitro Inhibition Assay: Plasma kallikrein inhibitory activity in vitro was determined using standard published methods (see e.g. Johansen et al., Int. J. Tiss. Reac. 1986, 8, 185; Shori et al., Biochem. Pharmacol., 1992, 43, 1209; Stiirzebecher et al., Biol. Chem. Hoppe-Seyler, 1992, 373, 1025). Human plasma kallikrein (Protogen) was incubated at 37° C. with the fluorogenic substrate H-DPro-Phe-Arg-AFC and various concentrations of the test compound. Residual enzyme activity (initial rate of reaction) was determined by measuring the change in optical absorbance at 410 nm and the IC50 value for the test compound was determined.
Measurement: IC50 = 91 nM (pChEMBL 7.039999961853027)
Descriptors: MW 596.7, ALogP 4.07, HBA 5, HBD 4, PSA 122.55, QED 0.17
Compound (SMILES): O=C1NC(=O)/C(=C/c2cnn3c(NC4CC4)cc(-c4ccc(C(=O)Nc5cccnc5)s4)nc23)N1
InChIKey: BUBFDSINQMUVNG-PXNMLYILSA-N
Target: Casein kinase II subunit beta (SINGLE PROTEIN), Homo sapiens
UniProt: P67870
Assay: Inhibition Assay: 5 uM ATP: Test compounds dissolved and diluted in DMSO (2 μl) were added to a reaction mixture comprising 10 μl of 5× Reaction Buffer (40 mM MOPS pH 7.0, 5 mM EDTA), 10 μl of recombinant human PIM2 solution (4 ng PIM-2 dissolved in dilution buffer (20 mM MOPS pH 7.0; EDTA 1 mM; 5% Glycerol; 0.01% Brij 35; 0.1%; 0.1% 2-mercaptoethanol; 1 mg/ml BSA)) and 8 ul of water. Reactions were initiated by the addition of 10 ul of ATP Solution (49% (15 mM MgCl2; 5 uM ATP) 1% ([γ-33P]ATP: Stock 1 mCi/100 μl; 3000 Ci/mmol (Perkin Elmer)) and 10 ul of substrate peptide solution (RSRSSYPAGT (SEQ ID NO.: 6), dissolved in water at a concentration of 1 mM), Reactions were maintained for 10 min at 30° C. The reactions were quenched with 100 ul of 0.75% Phosphoric acid, then transferred to and filtered through a Phosphocellulose filter plate (Millipore, MSPH-N6B-50). After washing each well 4 times with 0.75% Phosphoric acid, scintillation fluid (20 uL) was added to each well and
Measurement: IC50 > 2500 nM (pChEMBL 5.6020002365112305)
Descriptors: MW 486.52, ALogP 2.86, HBA 9, HBD 4, PSA 142.41, QED 0.24
Compound (SMILES): CN(C)C(=O)c1cc(C#N)ccc1C1C2=C(CCCC2=O)N(c2cccc(C(F)(F)F)c2)C(=O)N1C
InChIKey: OIVHHLLEYGZXCI-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 = 1.1 nM (pChEMBL 8.960000038146973)
Descriptors: MW 496.49, ALogP 4.9, HBA 4, HBD 0, PSA 84.72, QED 0.61
Compound (SMILES): Cc1cccc(N(C(=O)OC[C@H]2CC[C@@H](COCC(=O)O)CC2)c2ccccc2)c1
InChIKey: XEALUAITBGZBLD-BGYRXZFFSA-N
Target: Prostacyclin receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P43119
Assay: Agonist activity at human recombinant prostacyclin receptor expressed in CHO-K1 cells assessed as induction of cAMP accumulation incubated for 1 hr by HTRF assay
Measurement: EC50 = 2.7 nM (pChEMBL 8.569999694824219)
Descriptors: MW 411.5, ALogP 5.18, HBA 4, HBD 1, PSA 76.07, QED 0.65
Compound (SMILES): N#Cc1cc(-c2ccnc(Nc3ccnc(N4CCOCC4)c3)n2)cnc1N1CC[C@H](F)C1
InChIKey: NYMUSZKMHTWPPV-SFHVURJKSA-N
Target: Inhibitor of nuclear factor kappa-B kinase subunit epsilon (SINGLE PROTEIN), Homo sapiens
UniProt: Q14164
Assay: Time-Resolved Fluorescence-based Lanthascreen Assay: Kinase inhibition assays (10 μL) were performed at 20° C. in 384-well plate format. Compound IC50 values were determined at the apparent Km for ATP (20 μM) based on a radiometric assay (Invitrogen) using 8 or 10 point curves in duplicate. The final reaction conditions contained 400 nM fluorescein-IkBα substrate (DRHDSGLDSMKDE), 20 μM ATP, 2 nM or 8 nM IKKε or TBK1 kinase respectively, and 3% DMSO in kinase assay buffer consisting of 50 mM HEPES (pH 7.5), 10 mM MgCl, 1 mM EGTA, 0.01% Brij-35.Compound dilutions were prepared from 10 mM DMSO stocks by dilution into DMSO. Compound dilution series were further diluted in kinase assay buffer to give a 12% DMSO stock, the final concentration in the assay being 3% DMSO.The kinase phosphorylation assay was initiated by the addition of the kinase and the reaction was allowed to proceed for 1 h or 2.5 h for IKKε and TBK-1 kinase respectively.
Measurement: IC50 < 15 nM (pChEMBL 7.823999881744385)
Descriptors: MW 446.49, ALogP 2.93, HBA 9, HBD 1, PSA 103.09, QED 0.63
Compound (SMILES): CNC(=O)c1c(-c2ccc(F)cc2)oc2cc(N(CC(F)F)S(C)(=O)=O)c(-c3ccc4c(n3)-c3cc5c(F)cccc5n3CO4)cc12
InChIKey: VRIQKDGTKSMEEN-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1a (isolate H) (HCV)
UniProt: P27958
Assay: ELISA-Based Assay: Measurement of inhibition by compounds was performed using the HCV replicon system. Several different replicons encoding different HCV genotypes or mutations were used. In addition, potency measurements were made using different formats of the replicon assay, including different ways of measurements and different plating formats. See Jan M. Vrolijk et al., A replicons-based bioassay for the measurement of interferons in patients with chronic hepatitis C, 110 J. VIROLOGICAL METHODS 201 (2003); Steven S. Carroll et al., Inhibition of Hepatitis C Virus RNA Replication by 2'-Modified Nucleoside Analogs, 278(14) J. BIOLOGICAL CHEMISTRY 11979 (2003). However, the underlying principles are common to all of these determinations, and are outlined below.Stable neomycin phosphotransferase encoding replicons-harboring cell lines were used, so all cell lines were maintained under G418 selection prior to the assay. Potency was determined using a cell ELISA assay with an antibody to the replicons encoded NS3/4a protease. See Caterina Trozzi et al., In Vitro Selection and Characterization of Hepatitis C Virus Serine Protease Variants Resistant to an Active-Site Peptide Inhibitor, 77(6) J. Virol. 3669 (2003).
Measurement: IC50 = 8.4 nM (pChEMBL 8.079999923706055)
Descriptors: MW 664.64, ALogP 6.8, HBA 7, HBD 1, PSA 106.67, QED 0.19
Compound (SMILES): CSc1c(Cl)nc(-c2ccc(NC(=O)Nc3ccc(S(N)(=O)=O)cc3)cc2)nc1N1CCOCC1
InChIKey: ZKBZKPXYWXOMJD-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase mTOR (SINGLE PROTEIN), Homo sapiens
UniProt: P42345
Assay: mTOR Kinase Assay: The activity of the compound against mTOR was examined by measuring the incorporation of 33P from [γ-33P]-ATP into 4EBP1. His-tagged, recombinant human 4EBP1 was expressed in E. coli, purified by nickel-nitrilotriacetic acid (Ni-NTA) resins, and stored at −80 °C. Phosphorylation of 4EBP1 by mTOR was assayed in the presence or absence of the compound and performed in a final volume of 25 μl reaction buffer containing 300 ng 4EBP1, 50 ng recombinant mTOR (Invitrogen), 50 mM HEPES (pH 7.5), 1 mM EGTA, 0.01% Polysorbate 20, 10 mM MnCl2, 2.5 mM DTT, 10 μM ATP, and 0.5 μCi [Î³-33P]-ATP (PerkinElmir) for 30 min at 30 °C. The reactions were terminated by adding 3% phosphoric acid. The 33P labeled 4EBP1 was transferred onto UniFilter-96 GF/B plate (PerkinElmer) and quantified by Top Count Microplate Scintillation Counter (PerkinElmer). For primary screening of kinase activity inhibition, each test compound was evaluated at 10 M in duplicate.
Measurement: IC50 = 26.8 nM (pChEMBL 7.570000171661377)
Descriptors: MW 535.05, ALogP 3.65, HBA 8, HBD 3, PSA 139.54, QED 0.32
Compound (SMILES): O=C(NCCN1CCCC1=O)c1ccc(NC(=O)N2Cc3ccccc3C2)cc1
InChIKey: KJWUCMWEOOPUCH-UHFFFAOYSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Binding Assay : Probe 1: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-1251667.0. 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 = 35.4 nM (pChEMBL 7.449999809265137)
Descriptors: MW 392.46, ALogP 2.59, HBA 3, HBD 2, PSA 81.75, QED 0.82
Compound (SMILES): O=S(=O)(NCCN1CC2CN(CCc3ccc(F)cc3)CC(C1)O2)c1ccc(F)cc1
InChIKey: XODKOLJPPGQYSQ-UHFFFAOYSA-N
Target: Voltage-gated inwardly rectifying potassium channel KCNH2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q12809
Assay: Inhibition of human ERG expressed in CHO cells incubated for 10 mins after 3 hrs incubation with Rb+-load buffer by atomic absorption spectrometry based Rb+ efflux assay
Measurement: IC50 < 31622.78 nM (pChEMBL 4.5)
Descriptors: MW 451.54, ALogP 1.87, HBA 5, HBD 1, PSA 61.88, QED 0.66
Compound (SMILES): COc1ccccc1C(=O)c1ccc(C(=O)c2ccc(N(C)c3ccc(Cl)cc3)cn2)cc1C(=O)O
InChIKey: XOXQVJLXOZOINO-UHFFFAOYSA-N
Target: Leukotriene C4 synthase (SINGLE PROTEIN), Homo sapiens
UniProt: Q16873
Assay: Inhibition of recombinant human LTC4 synthase expressed in Pichia pastoris using LTA4 as substrate preincubated for 10 mins followed by substrate addition measured after 5 mins by HTRF method
Measurement: IC50 = 154 nM (pChEMBL 6.809999942779541)
Descriptors: MW 500.94, ALogP 5.67, HBA 6, HBD 1, PSA 96.8, QED 0.31
Compound (SMILES): COc1cccc2c1CCN(Cc1ccc([C@H](C)NC(=O)c3cn[nH]c3)cc1)C2
InChIKey: KOJAUBOTPMTGPW-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 = 184 nM (pChEMBL 6.739999771118164)
Descriptors: MW 390.49, ALogP 3.47, HBA 4, HBD 2, PSA 70.25, QED 0.68
Compound (SMILES): C[C@H]1C[C@H]2CSC(N)=N[C@@]2(c2nc(NC(=O)c3cn(C4CCC4)cn3)cs2)CO1
InChIKey: ITYWKJYCAPJVMB-ZKTNFTSUSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: BACE1 Cell-Free Assay: The substrate is Biotin-GLTNIKTEEISEISY^EVEFR-C[Oregon Green]KK-OH. The BACE1 enzyme is affinity purified material from conditioned media of CHO-K1 cells that have been transfected with a soluble BACE construct (BACE1deltaTM96His). Compounds are incubated in a ½ log dose response curve from a top concentration of 100 μM with BACE1 enzyme and the biotinylated fluorescent peptide in 384-well black plates (Thermo Scientific #4318). BACE1 is at a final concentration of 0.1 nM with a final concentration of peptide substrate of 150 nM in a reaction volume of 30 μL assay buffer [100 mM sodium acetate, pH 4.5 (brought to pH with acetic acid), and 0.001% Tween-20]. Plates are covered and incubated for 3 hours at 37 °C. The reaction is stopped with the addition of 30 μL of 1.5 μM Streptavidin (Pierce, #21125). After a 10 minute incubation at room temperature, plates are read on a PerkinElmer EnVision for fluorescence polarization (Ex485 nm/Em530 nm).
Measurement: IC50 = 34100 nM (pChEMBL 4.46999979019165)
Descriptors: MW 432.58, ALogP 3, HBA 9, HBD 2, PSA 107.42, QED 0.77
Compound (SMILES): CC1CC(N2CC(CC#N)(n3cc(-c4ncnc5[nH]ccc45)cn3)C2)CCN1C(=O)c1cccc(F)c1
InChIKey: UGNHTUCXHZUOAH-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: In Vitro JAK Kinase Assay: Compounds herein were tested for inhibitory activity of JAK targets according to the following in vitro assay described in Park et al., Analytical Biochemistry 1999, 269, 94-104. The catalytic domains of human JAK1 (a.a. 837-1142), Jak2 (a.a. 828-1132) and Jak3 (a.a. 781-1124) with an N-terminal His tag were expressed using baculovirus in insect cells and purified. The catalytic activity of JAK1, JAK2 or JAK3 was assayed by measuring the phosphorylation of a biotinylated peptide. The phosphorylated peptide was detected by homogenous time resolved fluorescence (HTRF). IC50s of compounds were measured for each kinase in the 40 microL reactions that contain the enzyme, ATP and 500 nM peptide in 50 mM Tris (pH 7.8) buffer with 100 mM NaCl, 5 mM DTT, and 0.1 mg/mL (0.01%) BSA. For the 1 mM IC50 measurements, ATP concentration in the reactions was 1 mM. Reactions were carried out at room temperature for 1 hr and then stopped with 20 μL 45 mM EDTA, 300 nM SA-APC, 6 nM Eu-Py20 in assay buffer (Perkin Elmer, Boston, Mass.). Binding to the Europium labeled antibody took place for 40 minutes and HTRF signal was measured on a Fusion plate reader (Perkin Elmer, Boston, Mass.).
Measurement: IC50 < 10 nM (pChEMBL 8)
Descriptors: MW 498.57, ALogP 3.58, HBA 7, HBD 1, PSA 106.73, QED 0.45
Compound (SMILES): Cn1cc(-c2cc(S(C)(=O)=O)ccc2OCCC2CC2)c2cc[nH]c2c1=O
InChIKey: WULUMURXICTAGJ-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 = 3.6 nM (pChEMBL 8.4399995803833)
Descriptors: MW 386.47, ALogP 3.12, HBA 5, HBD 1, PSA 81.16, QED 0.71
Compound (SMILES): C.CCCCN1CCN(C(=O)N2Cc3c(NC(=O)c4ccccn4)n[nH]c3C2(C)C)[C@@H](C)C1
InChIKey: RHIPLHOEQIBLAO-NTISSMGPSA-N
Target: Protein kinase C beta type (SINGLE PROTEIN), Homo sapiens
UniProt: P05771
Assay: Inhibition Assay: Protein Kinase C beta 2 (PKC beta II) catalyzes the production of ADP from ATP that accompanies the phosphoryl transfer to the PKC Pseudosubstrate peptide (A->S, RFARKGSLRQKNV). This transfer is coupled to the oxidation of p-NADH through the activities of Pyruvate Kinase (PK) and Lactate Dehydrogenase (LDH). (3-NADH conversion to NAD+ is monitored by the decrease in absorbance at 340 nm (e=6.22 cm-1 mM-1) using a Molecular Devices SPECTRA max PLUS spectrophotometer.A typical assay was carried out on a 96-well, clear microtiter plate in a Molecular Devices spectrophotometer for 20 minutes at 30 C. in 0.1 mL of assay buffer containing 50 mM HEPES, pH 7.4, 5 nM PKC, 23 units of pyruvate kinase, 33 units of lactate dehydrogenase, 0.15 mM peptide, 0.1 mM ATP, 1 mM DTT, 4 mM PEP, 8 mM MgCl2, 0.3 mM NADH, 60 mM CaCl2, 10 mg/mL PS, 50 ng/mL PMA, 7.5% DMSO and from about 10,000 nM to 0.169 nM compound inhibitor.
Measurement: Ki = 13.3 nM (pChEMBL 7.880000114440918)
Descriptors: MW 455.61
Compound (SMILES): O=C(c1ccc(Cl)cc1-n1nccn1)N1CCOC[C@H]1Cc1cc(-c2ncccn2)ccc1F
InChIKey: NNBZSPNSBCOMJO-LJQANCHMSA-N
Target: Orexin/Hypocretin receptor type 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O43613
Assay: OX Inhibition Assay: Chinese hamster ovary (CHO) cells expressing the human orexin-1 receptor and the human orexin-2 receptor, respectively, are grown in culture medium (Ham F-12 with L-Glutamine) containing 300 ug/mL G418, 100 U/mL penicillin, 100 ug/mL streptomycin and 10% heat inactivated fetal calf serum (FCS). The cells are seeded at 20,000 cells/well into 384-well black clear bottom sterile plates (Greiner). The seeded plates are incubated overnight at 37 C. in 5% CO2.Human orexin-A as an agonist is prepared as 1 mM stock solution in MeOH: water (1:1), diluted in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/L and 20 mM HEPES for use in the assay at a final concentration of 3 nM.Antagonists are prepared as 10 mM stock solution in DMSO, then diluted in 384-well plates using DMSO followed by a transfer of the dilutions into in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/L and 20 mM HEPES.
Measurement: IC50 = 3 nM (pChEMBL 8.520000457763672)
Descriptors: MW 478.92, ALogP 3.6, HBA 7, HBD 0, PSA 86.03, QED 0.44
Compound (SMILES): CC(C)(C)OC(=O)N1CCC(N(C(=O)c2ccc(-c3ccnnc3)cc2)C2CC2)CC1
InChIKey: CGVHJANETRZTCV-UHFFFAOYSA-N
Target: Glucose-dependent insulinotropic receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q8TDV5
Assay: AlphaScreen cAMP Assay: MIN6 cells [Miyazaki J et al. Endocrinology. 1990 July; 127(1):126-32] are stably transfected with an expression vector for human GPR119 cDNA (Acc. No. NP_848566). Min-6/hGPR119 cells are cultured in DMEM, 10% FBS, 50 uM β-mercaptoethanol, 0.3 mg/mL Geniticin, 2 mM GlutaMAX at 37° C. 5% CO2. For the assay, the cells are seeded in Optiplates (white, 384-well, 160W-barcoded, TC, sterile with lid, Cat. No. #6007688 (Perkin Elmer); 10000 cells/well; 50 ul). The plates covered with lids are then incubated for 24 hours at 37° C./5% CO2. After the medium is aspirated from the wells completely, 10 ul of the test compound are added, the compounds are diluted using stimulating buffer (140 mM NaCl, 3.6 mM KCl, 0.5 mM NaH2PO4, 0.5 mM MgSO4, 1.5 mM CaCl2, 10 mM Hepes, 5 mM NaHCO3; pH 7.4. 0.5 mM IBMX and 0.1% BSA, the final DMSO concentration is 1%). After 45 minutes incubation at room temperature (approx. 20° C.), the cAMP concentrations are determined using the AlphaScreen cAMP Assay Kit (Cat. No. #6760625R from PerkinElmer). 10 ul of Biotin-cAMP (final concentration 1 U/well in lysing buffer (5 mM Hepes (pH 7.4), 0.1% BSA, 0.5% Tween) and 10 uL Bead solution (final concentration 1 U/well in lysing buffer) are added. The plates are incubated for another 2 hours at room temperature. The cAMP concentrations are calculated using a cAMP standard curve from the Alpha Screen Counts. The data analysis is carried out by calculating the EC50 value and the maximum value based on a positive control, using suitable software (Graphpad Prism).
Measurement: EC50 = 236 nM (pChEMBL 6.630000114440918)
Descriptors: MW 422.53, ALogP 4.15, HBA 5, HBD 0, PSA 75.63, QED 0.74
Compound (SMILES): CC(C)(C)c1cc2cc(NC(=O)C3(c4ccc5c(c4)OCO5)CC3)c(F)cc2n1C[C@@H](O)CO
InChIKey: UUWCYYJATGFUAS-QGZVFWFLSA-N
Target: Cystic fibrosis transmembrane conductance regulator (SINGLE PROTEIN), Homo sapiens
UniProt: P13569
Assay: Correction activity at CFTR F508 deletion mutant (unknown origin) expressed in mouse NIH/3T3 cells assessed as increase in cAMP/genistein-dependent current amplitude incubated for 10 mins at 37 degC by patch clamp method
Measurement: EC50 = 65.95 nM (pChEMBL 7.179999828338623)
Descriptors: MW 468.53, ALogP 3.83, HBA 6, HBD 3, PSA 92.95, QED 0.51
Compound (SMILES): COc1cc(-c2cccc(C(F)(F)F)c2)ccc1-n1c(=O)cnc2cc(S(=O)(=O)Nc3ncccn3)ccc21
InChIKey: QFNUJUHXBXMVRI-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 = 78 nM (pChEMBL 7.110000133514404)
Descriptors: MW 553.52, ALogP 4.67, HBA 8, HBD 1, PSA 116.07, QED 0.32
Compound (SMILES): CCCCCCCn1ccc(C2(C)CC(c3ccc(C)cc3)=C(C#N)C(=O)N2)n1
InChIKey: MUEHTGWYFWABDR-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: MGAT LCMS Assay: The MGAT enzyme reactions were performed in CORNING Falcon 96-well Polypropylene plates, in a total volume of 60 uL of 50 mM Potassium Phosphate buffer pH 7.4, containing a final concentration of 100 uM 2-oleoylglycerol, 15 uM oleoyl-Coenzyme A and 0.0013 ug/uL Human or Mouse MGAT-2 or 0.0026 ug/uL 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 uL of ice cold methanol containing 1 ug/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, 50x2.1 mm column, was used for the chromatography of enzyme products.
Measurement: IC50 = 39 nM (pChEMBL 7.409999847412109)
Descriptors: MW 390.53, ALogP 4.87, HBA 4, HBD 1, PSA 70.71, QED 0.65
Compound (SMILES): CC(=O)N[C@@H](C)c1ccc(CN2CCc3c(ccc(OCC4(C(F)(F)F)CC4)c3Cl)C2)cc1
InChIKey: UOUVOFUMOUPJMC-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 = 55 nM (pChEMBL 7.260000228881836)
Descriptors: MW 480.96, ALogP 5.82, HBA 3, HBD 1, PSA 41.57, QED 0.54
Compound (SMILES): CN1Cc2c(C3=CCN(S(C)(=O)=O)CC3)[nH]c3nccc(c23)-c2cc(F)ccc21
InChIKey: BGYOGQNMBDFLOH-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: P50750
Assay: LANCE ULight TR-FRET Kinase Assay : CDK9 enzyme activities were measured using LANCE ULight TR-FRET kinase assay reagents (PerkinElmer, Waltham, Mass.). Compounds were diluted in 100% DMSO then diluted with 1:10 in serine/threonine kinase assay buffer containing 20 mM HEPES, 10 mM MgCl2, 100 mM Na3VO4, and 0.0075% Triton X-100. Equal volume of the compound dilutions was added to a final reaction mixture containing LANCE detection buffer (PerkinElmer CR97-100), 100 nM ULight MBP (PerkinElmer TRF0109M), 1000 μM ATP, and CDK9/Cyclin T1 (Carna Biosciences 04-110). The kinase reaction was carried out for 1 hour before addition of stopping buffer to a final of 20 mM EDTA and 0.5 nM of LANCE Ultra Europium anti-phospho-MBP antibody (PerkinElmer TRF0201M) in LANCE detection buffer. The reaction was incubated for 1 hour and the signal was read in Envision in TR-FRET mode (excitation at 320 nm and emission at 615/665 nm).
Measurement: IC50 = 25 nM (pChEMBL 7.599999904632568)
Descriptors: MW 412.49, ALogP 3.37, HBA 4, HBD 1, PSA 69.3, QED 0.7
Compound (SMILES): Cc1c(Cl)cccc1NC(=O)c1cc(NC(=O)c2ccccc2C(F)(F)F)cc2[nH]c([C@@H]3CCCN3C)nc12
InChIKey: QXNVHMFQJKUPJS-QHCPKHFHSA-N
Target: Prostaglandin E synthase (SINGLE PROTEIN), Homo sapiens
UniProt: O14684
Assay: Inhibition Assay: mPGES-1 microsome fractions were prepared from CHO-K1 cells transiently transfected with plasmid encoding the human mPGES-1cDNA. Microsomes were diluted with potassium phosphate buffer containing reduced glutathione (pH7.4), and DMSO containing test compound or DMSO alone was added (such that DMSO final concentration would be 1% in each) and incubated at 4° C. for 20 minutes. Then, the enzymatic reactions were initiated by the addition of PGH2 substrate (final concentration 1 μM) and incubated at 4° C. for 60 seconds. The reaction was terminated by the addition of a citrate solution (final citrate concentration 50 mM) containing ferric chloride (final concentration 1 mg/mL). PGE2 production in the enzyme reaction aliquot was measured using HTRF kit (Cisbio International, catalogue #62P2APEC).
Measurement: IC50 = 9.8 nM (pChEMBL 8.010000228881836)
Descriptors: MW 555.99, ALogP 6.81, HBA 4, HBD 3, PSA 90.12, QED 0.25
Compound (SMILES): O=C(C1CNC1)N1CCC(c2ccc(C(OCc3c(F)cccc3F)(C(F)(F)F)C(F)(F)F)cc2)(S(=O)(=O)c2ccc(F)cc2)C1
InChIKey: KIPALSCLAAICRP-UHFFFAOYSA-N
Target: Nuclear receptor ROR-gamma (SINGLE PROTEIN), Homo sapiens
UniProt: P51449
Assay: Scintillation Proximity Assay (SPA): The binding of potential ligands to RORγ is measured by competition with [3H] 25-hydroxycholesterol (Perkin Elmer NET674250UC) using a scintillation proximity assay (SPA) binding assay. The ligand binding domain of human RORγ (A262-S507) with an N-terminal His tag is expressed in E. coli and purified using nickel affinity chromatography. 15 ug/well RORγ (A262-S507) is incubated with test compound at varying concentrations in 3-fold serial dilution, with final concentrations ranging from 16.6 uM to 0.28 nM for 10 min at room temperature in PBS buffer (Invitrogen #14190-144) containing 0.5% fatty acid free BSA (Gemini Bio-Products, Cat. #700-107P) and 0.1% Glycerol (Sigma Cat#G5516). 10 nM of [3H] 25-hydroxycholesterol is then added, and the reaction is incubated for 10 min. 10 mg/mL of Copper-His Tag-PVT beads (Perkin Elmer cat #RPNQ0095) are added, and the mixture is incubated for 60 min. The reaction is read on a TopCount Microplate scintillation plate reader (Perkin Elmer). The competition data of the test compound over a range of concentrations was plotted as percentage inhibition of radioligand specifically bound in the absence of test compound (percent of total signal). After correcting for non-specific binding, IC50 values were determined. The IC50 value is defined as the concentration of test compound needed to reduce [3H] 25-hydroxycholesterol specific binding by 50% and is calculated using the four parameter logistic equation to fit the normalized data.
Measurement: IC50 = 1359 nM (pChEMBL 5.869999885559082)
Descriptors: MW 680.59, ALogP 5.76, HBA 5, HBD 1, PSA 75.71, QED 0.24
Compound (SMILES): CC(C)Oc1ccc(F)c(-c2c[nH]c3ccc(-c4nnc(N)o4)nc23)c1
InChIKey: USPNCTNCYQFTGT-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P11309
Assay: Enzyme Assay: The assay for the determination of Pim activity is based on the formation of phosphorylated biotinylated-BAD peptide at the Serine 112 residue (S112) and employs HTRF® (homogeneous time resolved fluorescence) technology to detect the product in a 96-well plate format. The phosphorylation of biotinylated-BAD (S112) peptide by full length recombinant Pim-1, Pim-2, or Pim-3 protein was detected with streptavidin:Allophycocyanin (APC) conjugate and a europium (Eu) labeled antibody directed against phosphorylated-BAD (S112). Excitation of Eu by a high energy laser light (337 nm) leads to a transfer of energy to the APC molecule, and results in an emission at 665 nm. The fluorescence is directly proportional to the amount of phosphorylated BAD peptide present in the reaction.Compounds were prepared in DMSO by conducting 3-fold serial dilutions to give a 10-point dosing curve having a high dose of 1 uM. A reference compound was included on each assay plate in order to validate that plate; on
Measurement: IC50 = 22.81 nM (pChEMBL 7.639999866485596)
Descriptors: MW 353.36, ALogP 3.79, HBA 6, HBD 2, PSA 102.85, QED 0.58