lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CN1CCC(NC(=O)CN2C(=O)N(c3cccc(C(F)(F)F)c3)C3=C(C(=O)CC3)C2c2ccc(C#N)cc2)CC1
InChIKey: PHHJHHJOBULGLY-UHFFFAOYSA-N
Target: Neutrophil elastase (SINGLE PROTEIN), Homo sapiens
UniProt: P08246
Assay: Neutrophil Elastase Assay: Materials: Human neutrophil elastase was purchased from Calbiochem (Cat. No.: 324681) and the elastase substrate MeOSuc-Ala-Ala-Pro-Val-AMC from Bachem (Cat. No.: I-1270). All other materials were of the highest grade commercially available. 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 uL of these compound dilutions were mixed with 10 ul 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 uL substrate solution in assay buffer were added (250 uM 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. Each plate contains wells with a high value control (DMSO+enzyme+substrate) and wells with a low value control (DMSO+inactivated enzyme+substrate). IC50 values were estimated using a sigmoidal concentration response curve with variable slope.
Measurement: IC50 = 22.6 nM (pChEMBL 7.650000095367432)
Descriptors: MW 551.57, ALogP 4.39, HBA 5, HBD 1, PSA 96.75, QED 0.59
Compound (SMILES): COC(=O)c1cnc(N2CCN(c3nnc(C(=O)c4ccccc4)c(C)c3C)C[C@H]2C)cn1
InChIKey: LWUAPVQWCGLGMN-OAHLLOKOSA-N
Target: Protein smoothened (SINGLE PROTEIN), Mus musculus
UniProt: P56726
Assay: Inhibition of mouse recombinant Shh protein in mouse TMHh12 cells assessed as inhibition of 1 nM Smo agonist-induced hedgehog signalling response by Gli-luciferase reporter gene assay
Measurement: IC50 = 100 nM (pChEMBL 7)
Descriptors: MW 446.51, ALogP 2.62, HBA 9, HBD 0, PSA 101.41, QED 0.43
Compound (SMILES): NC(=O)c1cccc2c1c(C[C@H](O)CO)cn2-c1ccc(Oc2ccc(F)cc2)cc1
InChIKey: WXUXQOGIRNAHNO-SFHVURJKSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: FLIPR Assay: Assay Buffer: The assay buffer was formulated by removing 120 mL from a 1 L bottle of fresh, sterile dH2O (Mediatech, Herndon, Va.) and adding 100 mL of 10×HBSS that does not contain Ca++ or Mg++ (Gibco, Invitrogen, Grand Island, N.Y.) followed by 20 mL of 1.0 M Hepes, pH 7.3 (Fisher Scientific, BP299-100). The final buffer consisted of 20 mM Hepes, pH 7.3, 1.261 mM CaCl2, 0.493 mM MgCl2, 0.407 mM Mg(SO)4, 5.33 mM KCl, 0.441 mM KH2PO4, 137 mM NaCl, 0.336 mM Na2HPO4 and 0.556 mM D-glucose (Hanks et al., Proc. Soc. Exp. Biol. Med. 71:196 (1949)), and the simple formulation was typically the basic buffer throughout the assay (i.e., all wash and addition steps).
Measurement: IC50 = 222 nM (pChEMBL 6.650000095367432)
Descriptors: MW 420.44, ALogP 3.56, HBA 5, HBD 3, PSA 97.71, QED 0.43
Compound (SMILES): NCCC[C@H]1CNC(=O)c2cc(-c3cccc(OC(F)(F)F)c3)c(-c3ccccc3)n21
InChIKey: YCKUAKXRPAUDMQ-KRWDZBQOSA-N
Target: Serine/threonine-protein kinase pim-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P11309
Assay: Trans-Phosphorylation Assay: The inhibitory activity of putative kinase inhibitors and the potency of selected compounds were determined using a trans-phosphorylation assay. Specific peptide or protein substrates were trans-phosphorylated by their specific ser-thr or tyr kinase in the presence of ATP traced with 33P-gamma-ATP, and in the presence of their own optimal buffer and cofactors.At the end of the phosphorylation reaction, more than 98% unlabeled ATP and radioactive ATP were captured by an excess of the ion exchange dowex resin; the resin then settled down to the bottom of the reaction plate by gravity. Supernatant was subsequently withdrawn and transferred into a counting plate, then evaluated by beta-counting. Kinase Buffer (KB):The buffer for PIM1 assay was composed of HEPES 50 mM, at pH 7.5, with 10 mM MgCl2, 1 mM DTT, 3 uM NaVO3, and 0.2 mg/mL BSA.Â  Full-length human PIM1 was expressed and purified as described in Bullock A N, et al., J. Biol. Chem. 2005, 280, 41675-82.
Measurement: IC50 = 19 nM (pChEMBL 7.71999979019165)
Descriptors: MW 429.44, ALogP 4.74, HBA 4, HBD 2, PSA 69.28, QED 0.59
Compound (SMILES): CC(C)(C)C(=O)N1CCN(C(=O)c2cc(Cc3n[nH]c(=O)c4c3NCCC4)ccc2F)CC1
InChIKey: PVRGCUYGIRVXMN-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 = 1.3 nM (pChEMBL 8.890000343322754)
Descriptors: MW 455.53, ALogP 2.19, HBA 5, HBD 2, PSA 98.4, QED 0.74
Compound (SMILES): Cc1cc(F)ccc1[C@H]1C[C@@]2(CCN1C(=O)N(C)[C@H](C)c1cc(C(F)(F)F)cc(C(F)(F)F)c1)CC(C(=O)[O-])CN2.[Na+]
InChIKey: OOIVQFSPQCKMRS-WYXRGQIUSA-M
Target: Substance-P receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P25103
Assay: Antagonist at human NK1 receptor expressed in recombinant BacMam virus infected human U2OS cells assessed as inhibition of substance P-induced intracellular calcium release preincubated for 30 mins followed by substance P addition by Fluo-4 calcium dye-based FLIPR assay
Measurement: Ki < 1 nM (pChEMBL 9)
Descriptors: MW 589.55, ALogP 6.55, HBA 3, HBD 2, PSA 72.88, QED 0.4
Compound (SMILES): CC(C)Oc1ccc(-c2nc(-c3cccc4c3CC[C@H]4NCC(=O)N3CC3)no2)cc1C#N
InChIKey: MKHXAWNHDCTILD-OAQYLSRUSA-N
Target: Sphingosine 1-phosphate receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21453
Assay: Agonist activity at human S1P1 expressed in CRE-bla CHOK1 cells assessed as inhibition of forskolin-induced cAMP formation preincubated for 4 hrs followed by FRET-based beta-lactamase fluorescent substrate addition measured after 2 hrs
Measurement: EC50 = 0.5 nM (pChEMBL 9.300000190734863)
Descriptors: MW 443.51, ALogP 3.48, HBA 7, HBD 1, PSA 104.05, QED 0.56
Compound (SMILES): Cc1nn(Cc2ccc(-c3cc4ccc(C(F)(F)F)cc4[nH]3)cc2C(F)(F)F)c(C)c1CC(=O)O
InChIKey: GGXRMVYEBQEOIG-UHFFFAOYSA-N
Target: Prostaglandin D2 receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Y4
Assay: Radioligand Binding Assay: The CRTH2 receptor binding assay is performed in a scintillation proximity assay (SPA) format with the radioligand [3H]-PGD2 (Perkin Elmer, NET616000MC). CHO-K1-hCRTH2 cell membranes are again homogenized by passing through a single use needle (Terumo, 23G) and diluted in SPA incubation buffer in suitable concentrations (0.5-10 ug protein/well). The SPA assay is set up in 96 well microtiter plates (Perkin Elmer, CatNo. 6005040) in SPA incubation buffer with a final volume of 200 ul per well and final concentration of 50 mM Tris-HCl, 10 mM MgCl2, 150 mM NaCl, 1 mM EDTA pH 7.4, 0.1% bovine serum albumin). The SPA assay mixture contains 60 ul of the membrane suspension, 80 ul of Wheat Germ Agglutinin coated PVT beads (GE Healthcare, RPNQ-0001, 0.3 mg/well), 40 ul of [3H]-PGD2 diluted in SPA buffer to a final concentration of 1 nM (50 000 dpm) and 20 ul of the test compound (dissolved in dimethylsulfoxid).
Measurement: Ki = 0.7 nM (pChEMBL 9.149999618530273)
Descriptors: MW 495.42, ALogP 6.36, HBA 3, HBD 2, PSA 70.91, QED 0.32
Compound (SMILES): C=CC(=O)Nc1cccc(Nc2nc(Nc3ccccc3OC)ncc2Cl)c1
InChIKey: IJBMVFJGJFRXOX-UHFFFAOYSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Omnia Assay: Briefly, 10x stocks of EGFR-WT (PV3872) from Invitrogen and EGFR-T790M/L858R (40350) from BPS Bioscience, San Diego, Calif., 1.13x ATP (AS001A) and appropriate Tyr-Sox conjugated peptide substrates (KCZ1001) were prepared in 1x kinase reaction buffer consisting of 20 mM Tris, pH 7.5, 5 mM MgCl2, 1 mM EGTA, 5 mM β-glycerophosphate, 5% glycerol (10x stock, KB002A) and 0.2 mM DTT (DS001A). 5 uL of each enzyme were pre-incubated in a Corning (#3574) 384-well, white, non-binding surface microtiter plate (Corning, N.Y.) for 30 min. at 25° C. with a 0.5 uL volume of 50% DMSO and serially diluted compounds prepared in 50% DMSO. Kinase reactions were started with the addition of 45 uL of the ATP/Tyr-Sox peptide substrate mix and monitored every 71 seconds for 60 minutes at λex360/λem485 in a Synergy4 plate reader from BioTek (Winooski, Vt.). At the conclusion of each assay, progress curves from each well were examined for linear reaction kinetics and fit statistics.
Measurement: IC50 = 650 nM (pChEMBL 6.190000057220459)
Descriptors: MW 395.85, ALogP 4.75, HBA 6, HBD 3, PSA 88.17, QED 0.5
Compound (SMILES): Cc1ncc(C(CNC(=O)C2COc3ncsc32)N2CCC(F)(F)CC2)cn1
InChIKey: MWNYPZLZUURAMT-UHFFFAOYSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: YO-PRO Assay: Cells were grown to confluency in adherent culture at 37° C. in a humidified 5% CO2 incubator (split 1/5 every 3-4 days with DMEM, 10% FCS, 1% Penicillin/Streptomycin, 250 μg/ml Geneticin). Adherent cells were detached by incubation with Trypsine (1 ml per 165 cm2 dish) for 2 minutes, then washed off with 10 ml PBS (without Mg2+ and Ca2+), and resuspended in DMEM, 10% FCS, 1% Penicillin/Streptomycin, no Geneticin. 10'000 cells per well (48 hours before the assay) or 25'000 cells per well (Vi-cell XR (Beckman Coulter) (24 hours before the assay) in 50 μl full medium were seeded on 384-well black-wall, clear bottom plates, that were coated before with 10 μl per well Poly-L-Lysine, incubated for 30-60 minutes at 37° C. and washed once with PBS. Medium was removed from cells and 50 μl of assay buffer containing 0.5 μM YO-PRO-1 was added into the wells. Solutions of antagonist compounds were prepared by serial dilutions of a 10 mM DMSO solution of the antagonist into PBS using a BioMek (Beckman Coulter). Each concentration was performed in duplicate. For IC50 measurements 10 concentration points were measured (10 μM being the highest concentration followed by 9 serial dilution steps 1/3). The cells were incubated with the antagonists of the present invention together with ATP at a final concentration of 250 μM for 90 minutes. During this time period, four time points were taken. Each time point comprised the average of several measurements made within a few seconds. Fluorescence was measured in the FLIPR tetra (Molecular Devices) using the filters appropriate for YO-PRO-1 fluorescence (excitation 485/20, emission 530/25).
Measurement: IC50 = 2035 nM (pChEMBL 5.690000057220459)
Descriptors: MW 409.46, ALogP 2.31, HBA 7, HBD 1, PSA 80.24, QED 0.82
Compound (SMILES): N#Cc1ccc(Oc2ccc3nc(C(=O)N[C@@H]4CCS(=O)(=O)C4)ccc3c2)cc1
InChIKey: NCQVEACMGYDASM-MRXNPFEDSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: FRET-Based Membrane Potential Assay: Two days prior to the experiment, frozen HEK293 cells stably expressing recombinant human Nav1.7 were quickly thawed and plated at 25,000 cells/well in growth medium [DMEM (Invitrogen #11965) with 10% HI FBS (Invitrogen #10082), 2 mM glutamine, 100 units/mL penicillin, 0.1 mg/mL streptomycin (PSG, Sigma #G1146), and 500 ug/mL Geneticin (Invitrogen #10131)] in black-walled, clear-bottom 384-well poly-D-lysine-coated assay plates (Greiner Bio-One, Frickenhausen, Germany) and incubated in a humidified 5% CO2 incubator at 37° C. On the day of the assay, medium was removed by aspiration, and cells were washed with assay buffer [HBSS (Invitrogen, Carlsbad, Calif.) containing 20 mM HEPES (Invitrogen, Carlsbad, Calif.)]. After washing, 30 uL assay buffer containing the fluorescent voltage-sensor probe CC2-DMPE (Invitrogen, Carlsbad, Calif.) at 20 uM and 0.01% pluronic F-127 (Invitrogen, Carlsbad, Calif.) was added to the cells. Cells were incubated for 40 minutes at room temperature in the dark. Following the incubation, the cells were washed and 30 uL assay buffer containing 2.5 uM DiSBAC2(3) substrate (Invitrogen, Carlsbad, Calif.) and 0.5 mM VABSC-1 (Invitrogen, Carlsbad, Calif.) was added to the cells. The cells were incubated for 90 minutes at room temperature in the dark. Fluorescence readings were made using a FLIPR TETRA (Molecular Devices, Sunnyvale Calif.) equipped with voltage-sensor probe optics. At the start of each experiment the optimal (EC80) concentration of depolarizing agent (veratridine) was determined by testing a dilution curve of assay buffer containing veratridine (Sigma-Aldrich, St. Louis, Mo.) and 1 mg/mL scorpion venom (SVqq, from Leiurus quinquestriatus; Sigma-Aldrich, St. Louis, Mo.). Compounds were dissolved in dimethyl sulfoxide, and 8-point, 1:3 dilution concentration-response curves were prepared in duplicate in dimethyl sulfoxide, followed by preparation of 0.8 uL/well daughter plates of the dilutions. Test compounds in the daughter plate were diluted to (Ë¿3x) solutions in assay buffer immediately before assaying. Using the FLIPR TETRA, 20 uL of the (3x) compound solutions were first added to the cells, then 20 uL of depolarizing solution (3xEC80 veratridine+SVqq) were added 3 minutes later to activate the channel. Changes in fluorescence were measured at wavelengths of 440-480 nm and 565-625 nm over the course of the experimental run. Membrane depolarization was expressed as a ratio of the maximum F440-480 nm/F565-625 nm reading above average baseline F440-480 nm/F565-625 nm reading. IC50 values were calculated from curve fits of the ratio data using a four-parameter logistic Hill equation (Accelrys Assay Explorer 3.3 Client, Accelrys, San Diego, Calif.) with percent inhibition plotted against compound concentration.
Measurement: IC50 = 710 nM (pChEMBL 6.150000095367432)
Descriptors: MW 407.45, ALogP 2.82, HBA 6, HBD 1, PSA 109.15, QED 0.71
Compound (SMILES): CN(c1ccc(C(=O)NCCc2c[nH]c3ccc(Cl)cc23)cc1)c1cccc(F)c1
InChIKey: NTDBHDKIUXTMBE-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 = 264 nM (pChEMBL 6.579999923706055)
Descriptors: MW 421.9, ALogP 5.7, HBA 2, HBD 2, PSA 48.13, QED 0.42
Compound (SMILES): Cc1cccc(C)c1N1CC(C(=O)Nc2cc(N3CCOCC3)cc(C(F)(F)F)c2)CC1=O
InChIKey: AMBDZUZGECLVGO-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 461.48, ALogP 4.15, HBA 4, HBD 1, PSA 61.88, QED 0.74
Compound (SMILES): CC(=O)c1cc2cnc(Nc3ccc(N4CCNCC4)cn3)nc2n(C2CCCC2)c1=O.Cl
InChIKey: UEKZXOFNQFAJMP-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 4/cyclin D1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P24385
Assay: Inhibition of Cdk4/Cyclin D (unknown origin) using retinoblastoma protein substrate and [gamma32P]ATP incubated for 15 mins by beta plate counter based assay
Measurement: IC50 = 123 nM (pChEMBL 6.909999847412109)
Descriptors: MW 433.52, ALogP 2.66, HBA 9, HBD 2, PSA 105.04, QED 0.59
Compound (SMILES): C(=C/c1ccccc1)\C(=N\Nc1nncc2ccccc12)c1ccccc1
InChIKey: IUZQUGLOCXBWCX-QCGZKTFGSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR-2 (unknown origin) using poly (Glu, Tyr) 4:1 as substrate after 1 hr by ELISA
Measurement: IC50 = 0.42 nM (pChEMBL 9.380000114440918)
Descriptors: MW 350.43, ALogP 5.16, HBA 4, HBD 1, PSA 50.17, QED 0.4
Compound (SMILES): COc1cc(-c2ccc(Cl)c(C)c2)ccc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3cnccn3)ccc21
InChIKey: XVDHIODRMYPUPH-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 = 35 nM (pChEMBL 7.460000038146973)
Descriptors: MW 533.01, ALogP 5.22, HBA 7, HBD 1, PSA 103.18, QED 0.32
Compound (SMILES): CC1(C)CCN(c2cccc(Nc3cc(-c4cccc(C(=O)O)c4)nn4ccnc34)n2)C1
InChIKey: IPHGZXOCVGUULA-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Inhibition Assay: Assay plates: 96-well MultiScreen 0.65 um filter plates (Millipore Cat. No.: MADVNOB10)Streptavidin coated beads: Streptavidin Sepharose, suspension 5.0 mL, in 50 mM EDTA/PBS diluted (1:100), (Amersham, Cat. No.: 17-5113-01)Compounds: 10 mM in 100% dimethylsulfoxide (DMSO), final conc.: compound 0.003-100 uM in 10% DMSOEnzyme: SYK RPA purified, truncated construct of Spleen Tyrosine Kinase aa 360-635, stock solution 1 mg/mL, MW: 31.2 KDa, final conc.:0.0005 uM. Peptide 1: biotinylated peptide is derived from a naturally occurring phosphor-acceptor consensus sequence (Biotin-EPEGDYEEVLE), special order from QCB, stock solution 20 mM, final conc.: 5.0 uM.ATP: Adenosine-5'-triphosphate 20 mM, (ROCHE Cat. No.: 93202720), final concentration: 20 uM Buffer: HEPES: 2-Hydroxyethyl piperazine-2-ethanesulfonic acid (Sigma, Cat. No.: H-3375)final concentration: 50 mM HEPES pH7.5BSA: Bovine Serum Albumin Fraction V, fatty acid free (Roche Diagnostics GmbH, Cat. No. 9100221).
Measurement: IC50 = 599.45 nM (pChEMBL 6.21999979019165)
Descriptors: MW 428.5, ALogP 4.47, HBA 7, HBD 2, PSA 95.65, QED 0.48
Compound (SMILES): CC(C)c1cccc(Nc2nc(Nc3cccc(OCC(=O)NCC(O)CO)c3)ncc2F)c1
InChIKey: GTISGSPRWHOXJD-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Biochemical Assay: Several 2,4-pyrimidinediamine compounds were tested for the ability to inhibit Syk kinase catalyzed phosphorylation of a peptide substrate in a biochemical fluorescenced polarization assay with isolated Syk kinase. In this experiment, Compounds were diluted to 1% DMSO in kinase buffer (20 mM HEPES, pH 7.4, 5 mM MgCl2, 2 mM MnCl2, 1 mM DTT, 0.1 mg/mL acetylated Bovine Gamma Globulin). Compound in 1% DMSO (0.2% DMSO final) was mixed with ATP/substrate solution at room temperature. Syk kinase (Upstate, Lake Placid N.Y.) was added to a final reaction volume of 20 uL, and the reaction was incubated for 30 minutes at room temperature. Final enzyme reaction conditions were 20 mM HEPES, pH 7.4, 5 mM MgCl2, 2 mM MnCl2, 1 mM DTT, 0.1 mg/mL acetylated Bovine Gamma Globulin, 0.125 ng Syk, 4 uM ATP, 2.5 uM peptide substrate (biotin-EQEDEPEGDYEEVLE-CONH2, SynPep Corporation). EDTA (10 mM final)/anti-phosphotyrosine antibody (1x final)/fluorescent phosphopeptide tracer (0.5x final) was added in FP Dilution Buffer to stop the reaction for a total volume of 40 uL according to manufacturer's instructions (PanVera Corporation) The plate was incubated for 30 minutes in the dark at room temperature. Plates were read on a Polarion fluorescence polarization plate reader (Tecan). Data were converted to amount of phosphopeptide present using a calibration curve generated by competition with the phosphopeptide competitor provided in the Tyrosine Kinase Assay Kit, Green (PanVera Corporation).
Measurement: IC50 = 187 nM (pChEMBL 6.730000019073486)
Descriptors: MW 469.52, ALogP 3.07, HBA 8, HBD 5, PSA 128.63, QED 0.29
Compound (SMILES): Cc1noc(C)c1-c1ccc2c(c1)C(c1ccccc1)(N1CCC[C@H](O)C1)C(=O)N2
InChIKey: UXFXOEGGFKVCDW-XGLRFROISA-N
Target: Bromodomain-containing protein 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P25440
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The assay was carried out in binding mixtures of the bromodomain protein, 0-10 μM compounds and tetra-acetylated histone peptide (SGRGAC.KGGAC.KGLGAC.KGGAAC.KRHGSGSK-biotin) in buffer containing 25 mM HEPES pH 7.5, 100 mM NaCl, 0.1% BSA, 0.05% CHAPS, and detection reagents. The detection reagents, including streptavidin-labeled Tb cryptate and XL665-labeled anti-6×His antibody, were added after binding equilibrium was achieved for protein, compound and peptide. Upon further incubation for 1 hr, the TR-FRET signals were recorded on a BMG PHERAstar FS instrument.
Measurement: IC50 = 9.6 nM (pChEMBL 8.020000457763672)
Descriptors: MW 403.48, ALogP 3.61, HBA 5, HBD 2, PSA 78.6, QED 0.7
Compound (SMILES): CCn1ncc(Cl)c1-c1cc(C(=O)N[C@H](CN)Cc2ccc(F)c(F)c2)oc1Cl
InChIKey: WBNREKJRGKMSSO-NSHDSACASA-N
Target: RAC-alpha serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P31749
Assay: Inhibition of full length human AKT1 expressed in Sf9 cells assessed as reduction in substrate phosphorylation using biotin-ahx-ARKRERAYSFGHHA-amide substrate and [gamma-33P]ATP incubated for 40 mins by top count microplate scintillation counting method
Measurement: IC50 <= 2.512 nM (pChEMBL 8.600000381469727)
Descriptors: MW 443.28, ALogP 4.05, HBA 5, HBD 2, PSA 86.08, QED 0.58
Compound (SMILES): CC1CC1C(=O)N1CCC(Oc2ccc(NC(=O)N3Cc4ccncc4C3)cc2)C1
InChIKey: GSXCQFDAXGUXIA-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 = 48.9 nM (pChEMBL 7.309999942779541)
Descriptors: MW 406.49, ALogP 3.27, HBA 4, HBD 1, PSA 74.77, QED 0.85
Compound (SMILES): CC(C)CN1C[C@@H]2CN(C(=O)Cn3cc(NC(=O)c4cnn5cccnc45)c(-c4cc(Cl)ccc4OC(F)F)n3)C[C@@H]2C1
InChIKey: XYQRBOLRNJJRSW-KDURUIRLSA-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 = 0.82 nM (pChEMBL 9.09000015258789)
Descriptors: MW 613.07, ALogP 4.15, HBA 9, HBD 1, PSA 109.89, QED 0.3
Compound (SMILES): NC(=O)c1ccc2[nH]nc(-c3cccc(OCCN4CCCCC4)c3)c2c1
InChIKey: QXGJVXWGUUKVOW-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 <= 1000 nM (pChEMBL 6)
Descriptors: MW 364.45, ALogP 3.19, HBA 4, HBD 2, PSA 84.24, QED 0.7
Compound (SMILES): C[C@H](Nc1ncc(F)c(N2C(=O)OC[C@@H]2[C@@H](C)O)n1)c1ccc(C(C)(C)O)cc1F
InChIKey: MGUKRKGLGOTAFE-LYOVBCGYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Fluorescence Biochemical Assay: The IDH1 (R132H) mutant catalyzes the reduced form of NADP+ (NADPH) and α-ketoglutarate (α-KG) to form nicotinamide adenine dinucleotide phosphate (NADP+) and R (−)-2-hydroxyglutarate (2HG). The reaction can be monitored kinetically by following the oxidation of NADPH to NADP+ which is measured using fluorescence, excitation at 355 nm and emission at 530 nm. Reactions were monitored using the Perkin-Elmer Envision, Model 2101. More specifically, the biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Cat. No. 781076) using a final reaction volume of 20 μL and the following assay buffer conditions: 50 mM HEPES pH 7.5, 10 mM MgCl2, 1 mM DTT, 0.02% BSA, 0.02% Tween-20, 10 μM NADPH and 100 μM α-KG. The final reaction mixture contained 2.5% DMSO and test compounds with concentrations ranging 0.0000008-25 μM. The IDH1 (R132H) enzyme was used at a final concentration of 10 nM.
Measurement: IC50 = 453 nM (pChEMBL 6.340000152587891)
Descriptors: MW 422.43, ALogP 2.86, HBA 7, HBD 3, PSA 107.81, QED 0.66
Compound (SMILES): Nc1ccc(-c2ccc3ncc4c(=O)[nH]c(=O)n(-c5cccc(C(F)(F)F)c5)c4c3n2)cn1
InChIKey: OIOOKLQHEDGJOE-UHFFFAOYSA-N
Target: Tyrosine-protein kinase ABL1 (SINGLE PROTEIN), Homo sapiens
UniProt: P00519
Assay: Mobility Shift Assay: Agents: 1-fold kinase buffer without MnCl2: 50 mM HEPES, pH 7.5, 0.0015% Brij-35, 10 mM MgCl2, 2 mM DTT. 1-fold kinase buffer with MnCl2: 50 mM HEPES, pH 7.5, 0.0015% Brij-35, 10 mM MgCl2, 10 mM MnCl2, 2 mM DTT. Termination solution: 100 mM HEPES, pH 7.5, 0.0015% Brij-35, 0.2% Coating Reagent#3, 50 mM EDTA. 2.5-fold kinase solution: The kinase was added to the 1-fold kinase buffer to form a 2.5-fold kinase solution. 2.5-fold substrate solution: FAM fluorescence-labelled polypeptide and ATP were added to the 1-fold kinase buffer to form a 2.5-fold substrate solution. Compounds: The final test concentration of the compound was 10 μM at maximum. Firstly, a 50-fold concentration (i.e. 500 μM) was prepared with 100% DMSO. The compound was diluted with 100% DMSO by 5 folds at 5 concentrations (successively 500 μM, 100 μM, 20 μM, 4 μM, and 0.8 μM). After diluting with 1-fold kinase buffer by 10 folds, 5 μl of the samples were transferred to a 384-well reaction plate. Two negative control wells and two positive control wells were set at each line respectively. Replication was made in a 96-well plate in five concentrations of 5-fold compound. 10% DMSO was added to the positive control well, and 5 μl of EDTA (250 mM) was added to the negative control well. Procedure: To the 384-well reaction plate was added 5-fold compound dissolved in 10% DMSO at 5 μl/well. To the 384-well reaction plate was added 2.5-fold kinase solution at 10 Î¼l/well. The plate was incubated at room temperature for 10 mins. To the 384-well reaction plate was added 2.5-fold substrate solution at 10 μl/well. The plate was incubated at 28° C. for a certain period. 25 μl termination solution was added to terminate the reaction.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 450.38, ALogP 3.29, HBA 7, HBD 2, PSA 119.55, QED 0.4
Compound (SMILES): C[C@H]1CN(c2ccc(C(C)(C)O)cn2)CCN1c1nnc(Cc2ccccc2Cl)c2c1CCC2
InChIKey: YVODGHXNTWUMSD-SFHVURJKSA-N
Target: Protein smoothened (SINGLE PROTEIN), Mus musculus
UniProt: P56726
Assay: Inhibition of mouse recombinant Shh protein in mouse TMHh12 cells assessed as inhibition of 1 nM Smo agonist-induced hedgehog signalling response by Gli-luciferase reporter gene assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 478.04, ALogP 4.55, HBA 6, HBD 1, PSA 65.38, QED 0.58
Compound (SMILES): O=C1NC(=O)/C(=C/c2cnn3c(NC4CC4)nc(NCc4ccc(F)c(Cl)c4)nc23)N1
InChIKey: NAAFTAWZXKBTBD-NSIKDUERSA-N
Target: Casein kinase II subunit beta (SINGLE PROTEIN), Homo sapiens
UniProt: P67870
Assay: Inhibition Assay: 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; 75 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 the resid
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 442.84, ALogP 2.28, HBA 8, HBD 4, PSA 125.34, QED 0.34
Compound (SMILES): Cc1nc2cccc(-c3cc4c([nH]3)CCNC4=O)c2nc1NC(C)(C)CS(C)(=O)=O
InChIKey: FGWLSBSYUGWQOF-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P11309
Assay: Homogeneous Time Resolved Fluorescence Assay (HTRF): 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 384-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 22-point dosing curve having a high dose of 1 μM. A reference compound was included on each assay plate [Costar 3658] in order to validate that plate; on one plate of every assay run, two additional reference compounds were included. The Reaction Buffer consisted of 45 mM Hepes, pH 7.0, 15 mM NaCl, and 1 mM MgCl. The quench/detection buffer consisted of 50 mM Tris, 100 mM NaCl, 0.05% BSA, 0.1% Tween and 3 mM EDTA. Biotinylated BAD peptide (Biopeptide), 10 mM ATP (Sigma), Labeled p-BAD (S112) mAb (Cell Signalling and Perkin Elmer) [with 0.05% BSA and 2 mM DTT added] streptavidin:Allophycocyanin [Perkin Elmer]. Final concentrations either Pim-1 enzyme [5 pM], or Pim-2 enzyme [0.5 pM], DMSO [1%], BLC BAD (S112) [0.5 μM], ATP [1.5 μM], streptavidin:Allophycocyanin [0.002 mg/mL] and biotinylated-BAD (S112) mAb [100 pM]. Initial incubations were carried out at RT (22° C.) for 30 min for both Pim-1 and for Pim-2. Pim enzyme is added to compound in buffer, and plates are incubated of 30 min. Biotinylated BAD and ATP are added and plates are incubated for 1 h. A mixture of labeled p-BAD (S112) mAb and quench/detection buffer are added and incubated for 2 h. Fluorescence was measured by an HTRF® Envision microplate reader.
Measurement: IC50 = 44.4 nM (pChEMBL 7.349999904632568)
Descriptors: MW 427.53, ALogP 2.45, HBA 6, HBD 3, PSA 116.84, QED 0.58
Compound (SMILES): CC(C)C[C@@H](c1c(O)c(C=O)c(O)c(C=O)c1O)[C@H]1CC[C@@H](C)[C@@]2(C)C1=CC[C@@H]1[C@H]2C1(C)C
InChIKey: ZZLZHWBNIIKVOG-YHPPXHETSA-N
Target: Tyrosine-protein phosphatase non-receptor type 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P18031
Assay: Inhibition of PTP1B (unknown origin) using pNPP as substrate after 3 mins by colorimetric analysis
Measurement: IC50 = 4300 nM (pChEMBL 5.369999885559082)
Descriptors: MW 454.61, ALogP 6.21, HBA 5, HBD 3, PSA 94.83, QED 0.35
Compound (SMILES): CC(C)n1cc(-c2ccnc(N)n2)c(-c2cc(Cl)cc(NS(C)(=O)=O)c2F)n1
InChIKey: XRCYSJXOEWSFPE-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition of B-Raf V600E mutant in human A375 cells harboring B-Raf V600E mutant assessed as reduction in cell proliferation incubated for 2 days by Bright-Glo luminescence assay
Measurement: IC50 = 69 nM (pChEMBL 7.159999847412109)
Descriptors: MW 424.89, ALogP 3.33, HBA 7, HBD 2, PSA 115.79, QED 0.65
Compound (SMILES): O=C1NC(=O)/C(=C/c2cnn3c(NC4CC4)nc(N4CCCC4)nc23)N1
InChIKey: ZSMDNUZIWPSKPU-XFFZJAGNSA-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 354.37, ALogP 0.48, HBA 8, HBD 3, PSA 116.55, QED 0.54
Compound (SMILES): O=C1NC(=O)/C(=C/c2cnn3c(NC4CC4)nc(NC4CCC(F)(F)CC4)nc23)N1
InChIKey: MHRQDUOISZBBDC-GHXNOFRVSA-N
Target: Casein kinase II subunit beta (SINGLE PROTEIN), Homo sapiens
UniProt: P67870
Assay: Inhibition Assay: 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; 75 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 the resid
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 418.41, ALogP 1.87, HBA 8, HBD 4, PSA 125.34, QED 0.43
Compound (SMILES): CNS1(O)C[C@@](C)(c2cc(NC(=O)c3ccc(F)cn3)ccc2F)NC(=N)N1C
InChIKey: LIAVENAQWXAHHF-SFHVURJKSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: HTRF FRET Assay: 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 3× the final desired concentration in a volume of 10 μl are preincubated with purified human BACE1 catalytic domain (3 nM in 10 μl) 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 μl of 600 nM QSY7-APPswe-Eu substrate (200 nM final) to give a final reaction volume of 30 μl 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.
Measurement: Ki = 10 nM (pChEMBL 8)
Descriptors: MW 424.48, ALogP 2.62, HBA 5, HBD 5, PSA 113.37, QED 0.52
Compound (SMILES): COc1cc(N2CCC(N(C)C)CC2)ccc1Nc1nccc(-c2c(-c3cccc(C(=O)Nc4c(F)cccc4F)c3)nc3ccccn23)n1
InChIKey: MIPKRMBRGKCCEI-UHFFFAOYSA-N
Target: Insulin-like growth factor 1 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08069
Assay: Inhibition of human GST-rTEV-tagged IGF1R (957 to 1367 residues) expressed in baculovirus infected insect Sf9 cells assessed as decrease in biotin-aminohexyl-AEEEEYMMMMAKKKK-NH2 phosphorylation after 1 hr by fluorometer
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 674.76, ALogP 7.27, HBA 9, HBD 2, PSA 99.92, QED 0.17
Compound (SMILES): COc1ccc(CC2CCCCN2C(=O)n2ncc(C(O)(c3ccc(F)cc3)c3ccc(F)cc3)n2)cc1
InChIKey: NEJATSOZXGUICW-UHFFFAOYSA-N
Target: Monoacylglycerol lipase ABHD6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9BV23
Assay: Inhibition of human ABHD6 using 2-AG as natural substrate by fluorescence based assay
Measurement: IC50 = 251.19 nM (pChEMBL 6.599999904632568)
Descriptors: MW 518.56, ALogP 4.91, HBA 6, HBD 1, PSA 80.48, QED 0.39
Compound (SMILES): CC[C@@H](NC(=O)c1cc(C(=O)N2CCC[C@@H]2C)n2c1COCC2)c1ccccc1
InChIKey: LLAVWQQOTBFOFF-QFBILLFUSA-N
Target: Voltage-gated inwardly rectifying potassium channel KCNH2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q12809
Assay: hERG Inhibition Assay: The effect of compounds of the formula I on the cloned human cardiac hERG channel was evaluated in an in vitro model using a whole-cell patch-clamping technique. CHO (Chinese hamster ovary) cells stably expressing hERG, the potassium channel underlying IKr currents in human hearts, were grown in HAM's F-12 media supplemented with 10% fetal bovine serum, 1x penicillin/streptomycin and 500 ug/ml G418 (Invitrogen, Carlsbad, Calif., USA) in an atmosphere of 95% air and 5% carbon dioxide. Cells used for patch-clamping were seeded on glass or plastic coverslips 12 to 36 hours before use. hERG channel currents were recorded at room temperature using the whole-cell configuration of the patch clamp technique with an Axopatch 200B amplifier (Axon Instruments, Foster City, Calif., USA). Briefly, electrodes (3 to 6 MΩ resistance) were fashioned from TW150F glass capillary tubes (World Precision Instruments, Sarasota, Fla., USA) and filled with pipette solution (containing 120 mM potassium aspartate, 20 mM potassium chloride, 4 mM adenosine triphosphate disodium salt, 5 mM HEPES, 1 mM magnesium chloride; pH adjusted to 7.2 with potassium hydroxide). hERG currents were initiated by a positive voltage pulse (20 mV) followed by a negative pulse (-40 mV) and were recorded for off-line analyses. Once hERG current from a cell perfused with external solution (containing 130 mM sodium chloride, 5 mM potassium chloride, 2.8 mM sodium acetate, 1 mM magnesium chloride, 10 mM HEPES, 10 mM glucose, 1 mM calcium chloride; pH adjusted to 7.4 with sodium hydroxide) without the test compound, i.e. control solution, was stabilized, the cell was perfused with external solution containing the test compound at specific concentrations.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 395.5, ALogP 3.52, HBA 4, HBD 1, PSA 63.57, QED 0.84
Compound (SMILES): O=S(=O)(c1ccccc1)N(Cc1cccc(OC(F)(F)F)c1)c1sc2ccccc2c1Br
InChIKey: IZFBRXHFLOQNJU-UHFFFAOYSA-N
Target: Transient receptor potential M8 protein (SINGLE PROTEIN), Canis lupus familiaris
UniProt: Q1A7N1
Assay: Functional Assay: The functional activity of compounds of the formula (I) was determined by measuring changes in intracellular calcium concentration using a Ca2+-sensitive fluorescent dye. The changes in fluorescent signal were monitored by a fluorescence plate reader, either a FLIPR (Molecular Devices) or FDSS (Hamamatsu). Increases in intracellular Ca2+ concentration were readily detected upon activation with icilin.At 24 hrs prior to assay, HEK293 cells stably expressing canine TRPM8 were seeded in culture medium in black wall, clear-base poly-D-lysine coated 384-well plates (BD Biosciences, NJ, USA) and grown overnight in 5% CO2 at 37° C. On assay day, growth media was removed and cells were loaded with Calcium 3 Dye (Molecular Devices) for 35 min at 37° C., under 5% CO2 and then for 25 min at room temperature and atmosphere. Subsequently, cells were tested for agonist-induced increases in intracellular Ca2+ levels using FLIPR or FDSS. Cells were challenged with a compound of the Formula (I) (at varying concentrations) and intracellular Ca2+ was measured for 5 min prior to the addition of icilin to all wells to achieve a final concentration that produces approximately an 80% maximal response. EC50 or IC50 values for compounds of the present invention were determined from eight-point dose-response studies.
Measurement: IC50 = 8.4 nM (pChEMBL 8.079999923706055)
Descriptors: MW 542.4, ALogP 6.96, HBA 4, HBD 0, PSA 46.61, QED 0.26
Compound (SMILES): CC(C)Oc1ccc(/N=c2\n(C)c(=O)n(CCOC3CCCCO3)c(=O)n2Cc2ccc(Cl)cc2)cc1F
InChIKey: NYQXKNHJLZHVCJ-QCWLDUFUSA-N
Target: P2X purinoceptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P56373
Assay: Inhibition Assay: Stably expressing cell line (C6BU-1 cell transfected with human P2X3 receptor gene (GenBank accession number Y07683)) was used. The cells were seeded in a 96-well microtiter plate at a concentration of 8000 cells/well and cultured in the medium (8.3% fetal bovine serum, 8.3% horse serum, 1% antibiotic and antifungal in DMEM) for one day at 37 C. under 5% carbon dioxide atmosphere. The medium was replaced with 4 uM Fluo-3-AM solution (pH 7.5) containing 20 mM HEPES, 137 mM NaCl, 2.7 mM KCl, 0.9 mM MgCl2, 5.0 mM CaCl2, 5.6 mM D-glucose, 2.5 mM probenecid, 10% BSA, and 0.08% Pluronic F-127, and incubated at 37° C. under 5% dioxide carbon atmosphere for one hour. The plate was washed with washing buffer (20 mM HEPES, 137 mM NaCl, 2.7 mM KCl, 0.9 mM MgCl2, 5.0 mM CaCl2, 5.6 mM D-glucose, 2.5 mM probenecid, pH 7.5), and each well was added with 40 uL of this buffer. The plate was placed in High-Throughput Screening System FDSS 3000 (Hamamatsu Photonics K.K.).
Measurement: IC50 = 459 nM (pChEMBL 6.340000152587891)
Descriptors: MW 547.03, ALogP 3.75, HBA 9, HBD 0, PSA 88.98, QED 0.41
Compound (SMILES): CC(=O)NCc1nc2c(-c3cnc[nH]3)cc(N3CCOCC3)nn2c1Cc1cccc(C(F)(F)F)c1C
InChIKey: GLORCTPRDHXJPK-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42336
Assay: Binding affinity to human PI3Kalpha (108 to 1068 residues) expressed in mammalian expression system by KINOMEscan assay
Measurement: Kd > 30200 nM (pChEMBL 4.519999980926514)
Descriptors: MW 513.52, ALogP 3.51, HBA 7, HBD 2, PSA 100.44, QED 0.41
Compound (SMILES): COCCOC(=O)N[C@H]1CC[C@H](Nc2nc(NC3CC3)n3ncc(/C=C4\NC(=O)NC4=O)c3n2)CC1
InChIKey: MXBPDWFIVWCZNK-JEHBBDIFSA-N
Target: Casein kinase II subunit beta (SINGLE PROTEIN), Homo sapiens
UniProt: P67870
Assay: Inhibition Assay: 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; 75 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 the resid
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 499.53, ALogP 0.97, HBA 11, HBD 5, PSA 172.9, QED 0.19
Compound (SMILES): O=C1CCc2c(cccc2OCCCN2CCN(c3cccc4sccc34)CC2)N1
InChIKey: TZHCOSMLWPDCNG-UHFFFAOYSA-N
Target: D(2) dopamine receptor (SINGLE PROTEIN), Rattus norvegicus
UniProt: P61169
Assay: Dopamine D2 Receptor Binding Assay: The assay was performed according to the method by Kohler et al. (Kohler C, Hall H, Ogren S O and Gawell L, Specific in vitro and in vivo binding of 3H-raclopride. A potent substituted benzamide drug with high affinity for dopamine D-2 receptors in the rat brain. Biochem. Pharmacol., 1985; 34: 2251-2259). The binding assay was performed using 40 ul of the membrane specimen, 20 ul of [3H]-raclopride (final concentration 1 to 2 nM), 20 ul of a test drug and 50 mM Tris-hydrochloric acid buffer (containing 120 mM NaCl, 5 mM KCl, 2 mM CaCl2, 1 mM MgCl2, pH 7.4) so that the total amount was 200 ul (final dimethylsulfoxide concentration 1%). The reaction was performed at room temperature for 1 hour and terminated by conducting suction filtration with a cell harvester on a glass fiber filter plate. The filter plate made of glass fiber was washed with 50 mM Tris-hydrochloric acid buffer (pH 7.4), and after dried, a microplate liquid scintillation cocktail was added.
Measurement: Ki = 3.3 nM (pChEMBL 8.479999542236328)
Descriptors: MW 421.57, ALogP 4.38, HBA 5, HBD 1, PSA 44.81, QED 0.6