lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CC(=O)N1CCN(CC2CCC(NC(=O)c3cc4c(C)nn(C5CCOCC5)c4s3)CC2)CC1
InChIKey: STBGSMKSTHGKRX-UHFFFAOYSA-N
Target: Phosphodiesterase 4B (SINGLE PROTEIN), Mus musculus
UniProt: Q8VBU5
Assay: Inhibition Assay: The PDE 4 (phosphodiesterase IV) inhibiting effect of the compounds of the present invention was performed by the following method, which was modified assay method described in Biochemical. Pharmacol. 48(6), 1219-1223 (1994).  (1) The active fraction of PDE 4 (phosphodiesterase IV) was obtained. That is, the livers obtained from three Balb/c mice (male, 12 weeks: obtainable from CLEA Japan, Inc.) were suspended with 30 mL of buffer solution B [20 mM of bis-tris, 5 mM of 2-mercaptoethnol, 2 mM of benzamidine, 2 mM of EDTA, 0.1 mM of 4-(2-aminoethyl)benzensulfonyl hydrochloride, 50 mM of sodium acetate; pH 6.5], then homogenized by Polytron homogenizer. The homogenate were centrifuged under 25,000xG for 10 minutes at 4 C. The supernatant was separated and thus obtained supernatant was further centrifuged under 100,000xG for 60 minutes at 4Â° C., and then filtrated with 0.2 um filter to obtain the soluble fraction.
Measurement: IC50 = 7900 nM (pChEMBL 5.099999904632568)
Descriptors: MW 487.67, ALogP 3.21, HBA 7, HBD 1, PSA 79.7, QED 0.7
Compound (SMILES): CCOCc1c(-c2cc3cc(C)cc(OC)c3s2)c2c(N)ncnn2c1C(=O)N(C)C
InChIKey: IZEWFDKAZGEPEK-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Kinase Assay: For the assay, 50 nl of each stock solution of the test compound in DMSO was pipetted into a black, low-volume 384-well microtiter plate (Greiner Bio-One, Frickenhausen, Germany) 2 ul of a solution of the above FGFR-3 fusion protein in aqueous assay buffer [8 mM MOPS pH 7.0, 10 mM magnesium acetate, 1.0 mM dithiothreitol, 0.05% (w/v) bovine serum albumin (BSA), 0.07% (v/v) Tween-20, 0.2 mM EDTA] was added, and the mixture was incubated for 15 min at 22° C. to allow pre-binding of the test compound to the enzyme. Then, the kinase reaction was started by the addition of 3 ul of a solution of adenosine triphosphate (ATP, 16.7 uM; final concentration in the 5 ul assay volume=10 uM) and substrate (0.8 uM; final concentration in the 5 ul assay volume=0.5 uM) in assay buffer, and the resulting mixture was incubated for a reaction time of 60 min at 22° C. The concentration of FGFR-3 fusion protein was adjusted depending on the activity of the enzyme lot.
Measurement: IC50 = 9.6 nM (pChEMBL 8.020000457763672)
Descriptors: MW 439.54, ALogP 3.75, HBA 8, HBD 1, PSA 94.98, QED 0.49
Compound (SMILES): CNCCN1CCC(Cc2ccccc2)C1
InChIKey: VWJBIFGWCPEUEE-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q96LA8
Assay: Inhibition of human full length PRMT6 (1 to 375 residues) expressed in baculovirus expression system assessed as inhibition of methylation activity preincubated with protein followed by addition of biotin labeled histone H4 (1 to 24) peptide as substrate and [3H]-SAM measured after 3 hrs by scintillation proximity assay
Measurement: IC50 = 70 nM (pChEMBL 7.159999847412109)
Descriptors: MW 218.34, ALogP 1.77, HBA 2, HBD 1, PSA 15.27, QED 0.81
Compound (SMILES): CNC(=O)c1ccc(N2CCN(S(=O)(=O)c3ccc(OC(C)C)cc3)CC2)cc1OCC(=O)O
InChIKey: FHZDWXSQNKRBSI-UHFFFAOYSA-N
Target: Prostaglandin D2 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q13258
Assay: Displacement of [3H]-PGD2 from PGD2 receptor in human platelet membranes after 2 hrs by micro beta scintillation counting analysis
Measurement: Ki = 5000 nM (pChEMBL 5.300000190734863)
Descriptors: MW 491.57, ALogP 1.81, HBA 7, HBD 2, PSA 125.48, QED 0.54
Compound (SMILES): COc1ccc2nc(NC3=NC[C@]4(CN5CCC4CC5)O3)sc2c1
InChIKey: KIDXXFXVPLATJJ-QGZVFWFLSA-N
Target: 5-hydroxytryptamine receptor 3A (SINGLE PROTEIN), Homo sapiens
UniProt: P46098
Assay: Antagonist activity at human 5-HT3A expressed in HEK293 cells assessed as reduction in acetylcholine-induced activity preincubated for 30 mins followed by addition of acetylcholine measured for 2 mins by Fluo-4AM dye based FLIPR assay
Measurement: IC50 > 97000 nM (pChEMBL 4.013000011444092)
Descriptors: MW 344.44, ALogP 2.57, HBA 7, HBD 1, PSA 58.98, QED 0.91
Compound (SMILES): Cc1noc(C)c1C(=O)N1CC=C(c2ccc(NC(=O)N3Cc4ccccc4C3)cc2)CC1
InChIKey: XZDWISPPWUOXKB-UHFFFAOYSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Binding Assay : Probe 2:The assay was carried out in 18 uL low volume plates (Owens Corning) in reaction buffer (50 mM HEPES (NaOH), pH 7.5, 100 mM NaCl, 10 mM MgCl2, 1 mM DTT, 1% Glycerol) using 6.8 nM recombinant, human, C-terminally-His tagged NAMPT, 1 nM Tb-anti-His antibody (Invitrogen, Cat #PV5895), and 200 nM probe (Oregon Green 488-conjugated APO0866; A-1287128.0. Plates were covered, and reactions were carried out for 2-3 hours. Plates were read with Envision (Laser Lantha low volume protocol) after 2 to 3 hours. Excitation was carried out at 337 nm, and the ratio of emission of Oregon Green (520 nm) to terbium (492 nm) was determined and used to calculate IC50 values of test compounds.
Measurement: IC50 = 11.8 nM (pChEMBL 7.929999828338623)
Descriptors: MW 442.52, ALogP 4.77, HBA 4, HBD 1, PSA 78.68, QED 0.63
Compound (SMILES): CNc1noc2ccc(-c3nn(C(C)C)c4ncnc(N)c34)cc12
InChIKey: TWWYHXMMESUYNP-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42336
Assay: Inhibition of human PI3Kalpha using PIP2 as substrate by HTRF assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 323.36, ALogP 2.84, HBA 8, HBD 2, PSA 107.68, QED 0.6
Compound (SMILES): CC1(C)CC(=O)C2=C(C1)N(c1ccc(S(N)(=O)=O)cc1)C1=C(C(=O)CC(C)(C)C1)C2c1ccc(-c2nnn[nH]2)cc1
InChIKey: XHXARUOKKJEWLI-UHFFFAOYSA-N
Target: Carbonic anhydrase 7 (SINGLE PROTEIN), Homo sapiens
UniProt: P43166
Assay: Inhibition of human recombinant carbonic anhydrase 7 expressed in Escherichia coli assessed as reduction in CO2 hydration preincubated for 15 mins prior to testing measured for 10 to 100 secs by stopped-flow assay
Measurement: Ki = 45.3 nM (pChEMBL 7.340000152587891)
Descriptors: MW 572.69, ALogP 4.4, HBA 8, HBD 2, PSA 152, QED 0.47
Compound (SMILES): Fc1ccc(CN2CCC(c3nnn4cnc5[nH]ccc5c34)CC2)nc1
InChIKey: NDKFWXSXVFIJIN-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 200 nM (pChEMBL 6.699999809265137)
Descriptors: MW 351.39, ALogP 2.52, HBA 6, HBD 1, PSA 75, QED 0.61
Compound (SMILES): O=c1cc(NC2CCN(S(=O)(=O)C(F)(F)F)CC2)c2cc(C(O)(c3ccc(F)cc3)c3ccc(Cl)cc3)ccc2[nH]1
InChIKey: UYAUGGNRWFCFDZ-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 = 9700 nM (pChEMBL 5.010000228881836)
Descriptors: MW 610.03, ALogP 5.33, HBA 5, HBD 3, PSA 102.5, QED 0.2
Compound (SMILES): COc1cc(OC)cc(N(CCN2CCCC2)c2ccc3ncc(-c4cn[nH]c4)nc3c2)c1.Cl
InChIKey: BJTAMMVWXSCQFA-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 444.54, ALogP 4.27, HBA 7, HBD 1, PSA 79.4, QED 0.43
Compound (SMILES): CCC[C@H](Nc1nc(N)nc(N)c1Cl)c1nc2ccc(F)c(Cl)c2c(=O)n1-c1ccc(N)nc1
InChIKey: YMBSDVFEBBJZGE-LBPRGKRZSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit beta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42338
Assay: Time-Resolved Fluorescence Resonance Energy transfer (TR-FRET) Assay: Class I PI3K isoforms were expressed and purified as heterodimeric recombinant proteins. All assay reagents and buffers for the TR-FRET assay were purchased from Millipore. PI3K isoforms were assayed under initial rate conditions in the presence of 25 mM Hepes (pH 7.4), and 2× Km ATP (75-500 μM), 2 μM PIP2, 5% glycerol, 5 mM MgCl2, 50 mM NaCl, 0.05% (v/v) Chaps, 1 mM dithiothreitol, 1% (v/v) DMSO at the following concentrations for each isoforms: PI3Kα, PI3Kβ, and PI3Kδ between 25 and 50 pM, and PI3Kγ at 2 nM. After an assay reaction time of 30 minutes at 25° C., reactions were terminated with a final concentration of 10 mM EDTA, 10 nM labeled-PIP3, and 35 nM Europium labeled GRP-1 detector protein before reading TR-FRET on an Envision plate reader (Ex: 340 nm; Em: 615/665 nm; 10lps delay and 500 μs read window).
Measurement: IC50 = 530 nM (pChEMBL 6.28000020980835)
Descriptors: MW 504.36, ALogP 3.72, HBA 10, HBD 4, PSA 163.65, QED 0.31
Compound (SMILES): COc1cccc2c1C=[N+](c1ccccc1)CC2.[Br-]
InChIKey: TWVQBZQJADPFBR-UHFFFAOYSA-M
Target: Cholinesterase (SINGLE PROTEIN), Homo sapiens
UniProt: P06276
Assay: Inhibition of BuChE (unknown origin) preincubated for 30 mins followed by substrate addition acetylthiocholineiodide measured after 40 mins by Ellman's method
Measurement: IC50 = 35400 nM (pChEMBL 4.449999809265137)
Descriptors: MW 238.31, ALogP 3.01, HBA 1, HBD 0, PSA 12.24, QED 0.73
Compound (SMILES): C[C@@H]1CN(Cc2ccc(C(F)(F)F)o2)c2ccccc2[C@H](C)N1C(=O)NCc1ccccc1
InChIKey: SYDUPIQUDQXBIQ-MSOLQXFVSA-N
Target: Voltage-dependent T-type calcium channel subunit alpha-1H (SINGLE PROTEIN), Homo sapiens
UniProt: O95180
Assay: Channel blocking activity at recombinant human CaV 3.2 channel expressed in HEK293 cells assessed as inhibition of Ca2+ flux preincubated for 3 mins followed by CaCl2 addition by fluo-4 dye-based FLIPR assay
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 457.5, ALogP 5.98, HBA 3, HBD 1, PSA 48.72, QED 0.52
Compound (SMILES): COc1ccc(Cl)c(N(CCNC(C)C)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1.Cl
InChIKey: QIFORRQQZPOUPH-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 7079.46 nM (pChEMBL 5.150000095367432)
Descriptors: MW 450.97, ALogP 4.83, HBA 7, HBD 1, PSA 68.1, QED 0.41
Compound (SMILES): CC(C)(C)c1cc(NC(=O)Nc2ccc(Oc3ccnc(Nc4ccc5c(c4)NC(=O)C5(C)C)n3)c3ccccc23)n(-c2ccc(OCCN3CCOCC3)cc2)n1
InChIKey: SALBBGQOFCZHKF-UHFFFAOYSA-N
Target: Proto-oncogene tyrosine-protein kinase Src (SINGLE PROTEIN), Homo sapiens
UniProt: P12931
Assay: c-Src and Syk Enzyme Inhibition Assay: The inhibitory activities of compounds of the invention against c-Src and Syk enzymes (Invitrogen), are evaluated in a similar fashion to that described hereinabove. The relevant enzyme (3000 ng/mL or 2000 ng/mL respectively, 2.5 uL) is incubated with the test compound (either 4 ug/mL, 0.4 ug/mL, 0.04 ug/mL, or 0.004 ug/mL, 2.5 uL each) for 2 hr at RT. The FRET peptides (8 uM, 2.5 uL), and appropriate ATP solutions (2.5 uL, 800 uM for c-Src, and 60 uM ATP for Syk) are then added to the enzymes/compound mixtures and incubated for 1 hr. Development reagent (protease, 5 uL) is added for 1 hr prior to detection in a fluorescence microplate reader (Varioskan Flash, ThermoFisher Scientific).
Measurement: IC50 = 5 nM (pChEMBL 8.300000190734863)
Descriptors: MW 781.92, ALogP 8.23, HBA 11, HBD 4, PSA 156.79, QED 0.1
Compound (SMILES): COc1ccnc(OC)c1C1CCCC(=O)N1Cc1cccc(-c2csc(C)n2)c1
InChIKey: ILPKRFOPCQBPFG-UHFFFAOYSA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: In Vitro Assay: Intracellular Calcium Measurements : In Vitro Assay: Intracellular Calcium Measurements: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 g/ml G418, 100 U/ml penicillin, 100 g/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. On the day of the assay, 50 μl of staining buffer (HBSS containing 1% FCS, 20 mM HEPES, NaHCO3: 0.375 g/l, 5 mM probenecid (Sigma) and 3 M of the fluorescent calcium indicator fluo-4 AM (1 mM stock solution in DMSO, containing 10% pluronic) is added to each well. The 384-well cell-plates are incubated for 50 min at 37° C. in 5% CO2 followed by equilibration at RT for 30 min before measurement.
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 423.54, ALogP 4.78, HBA 6, HBD 0, PSA 64.55, QED 0.57
Compound (SMILES): N#Cc1nc(-c2ccc(OCCC3CCNCC3)c(C(F)(F)F)c2)c2ccn(CC3CCCO3)c2n1
InChIKey: DKGCAUXMTIITRY-UHFFFAOYSA-N
Target: Cathepsin S (SINGLE PROTEIN), Mus musculus
UniProt: O70370
Assay: Inhibition Assay: First, spleen cells collected from C57BL/6 mice were seeded in a 96-well plate so as to be a 1× 105 cells/well, then, 5-fold dilution of 10 mM DMSO solution of the test compound was serially performed with RPMI 1640 medium (including 10% fetal bovine serum (FCS), 5×10^−5 M 2-mercaptoethanol, 50 IU/mL penicillin and 50 μg/mL streptomycin) nine times such that the final concentration becomes 0.026 nM to 10 μM, and the resultant products were added thereto (the final DMSO concentration was 0.1%). At the same time, LPS (Sigma L4005) was added to the wells such that the final concentration becomes 2 μg/mL, and culture was performed at 37° C. for 48 hours in 5% CO2. After culturing, the cells were stained at 4° C. for 20 minutes with PE-labeled streptavidin (BD BIOSCIENCE 554061) and biotin-labeled YAe antibody (EBIOSCIENCE 13-5741-85) together with FITC-labeled anti-mouse B220 antibody (BD BIOSCIENCE 553088), and an expression level (fluorescence intensity of YAe-bioti
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 499.54, ALogP 4.94, HBA 7, HBD 1, PSA 84.99, QED 0.5
Compound (SMILES): O=C1/C(=C/c2ccc(=O)n(O)c2)Oc2cc(O)cc(O)c21
InChIKey: PVVQBJMLBJRPGS-JYOAFUTRSA-N
Target: Tyrosinase (SINGLE PROTEIN), Homo sapiens
UniProt: P14679
Assay: Inhibition of recombinant human tyrosinase expressed in baculovirus infected Sf9 cells using L-DOPA as substrate by spectrophotometric analysis
Measurement: IC50 = 30000 nM (pChEMBL 4.519999980926514)
Descriptors: MW 287.23, ALogP 1.11, HBA 7, HBD 3, PSA 108.99, QED 0.54
Compound (SMILES): c1cn2nc(NC3=NC[C@@]4(CN5CCC4CC5)O3)ccc2n1
InChIKey: LUJKACIVKPRMBY-HNNXBMFYSA-N
Target: 5-hydroxytryptamine receptor 3A (SINGLE PROTEIN), Homo sapiens
UniProt: P46098
Assay: Antagonist activity at 5-HT3A receptor (unknown origin) expressed in HEK293 cells assessed as inhibition of Ca2+ flux by Fluo-4-AM dye based FLIPR assay
Measurement: IC50 = 250 nM (pChEMBL 6.599999904632568)
Descriptors: MW 298.35, ALogP 0.99, HBA 7, HBD 1, PSA 67.05, QED 0.85
Compound (SMILES): O=c1cc(/C=C/c2ccc(C(F)(F)F)cc2)c2cc(C(c3ccccc3)c3ccc(Cl)cc3)ccc2[nH]1
InChIKey: RBYRKKSDIDNREM-RMKNXTFCSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 515.96, ALogP 8.55, HBA 1, HBD 1, PSA 32.86, QED 0.23
Compound (SMILES): COc1cccc(OC)c1C1CCCC(=O)N1C(C)c1cnc(OC(F)F)c(Cl)c1
InChIKey: HMIGQZCCCBMNIM-UHFFFAOYSA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: In Vitro Assay: Intracellular Calcium Measurements : In Vitro Assay: Intracellular Calcium Measurements: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 g/ml G418, 100 U/ml penicillin, 100 g/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. On the day of the assay, 50 μl of staining buffer (HBSS containing 1% FCS, 20 mM HEPES, NaHCO3: 0.375 g/l, 5 mM probenecid (Sigma) and 3 M of the fluorescent calcium indicator fluo-4 AM (1 mM stock solution in DMSO, containing 10% pluronic) is added to each well. The 384-well cell-plates are incubated for 50 min at 37° C. in 5% CO2 followed by equilibration at RT for 30 min before measurement.
Measurement: IC50 = 34 nM (pChEMBL 7.46999979019165)
Descriptors: MW 440.87, ALogP 5.17, HBA 5, HBD 0, PSA 60.89, QED 0.59
Compound (SMILES): Clc1cc(Br)ccc1Nc1ncc(Br)c(Nc2cc(C3CC3)[nH]n2)n1
InChIKey: NCAWMSOMZBQWRC-UHFFFAOYSA-N
Target: Insulin-like growth factor 1 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08069
Assay: Inhibition of human IGF1R kinase domain (M954 to C1367 residues) tri-phosphorylated enzyme form expressed in baculovirus based expression system using poly-EY peptide incubated for 2 hrs by luciferase-luciferin coupled chemiluminescent assay
Measurement: IC50 = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 484.58, ALogP 5.74, HBA 5, HBD 3, PSA 78.52, QED 0.43
Compound (SMILES): Cc1cc2c(-c3cnc(N)c(C(=O)Nc4ncccc4N4CCC(N)CC4)n3)cccc2[nH]1
InChIKey: AODINEOJAWVRMB-UHFFFAOYSA-N
Target: Protein kinase C theta type (SINGLE PROTEIN), Homo sapiens
UniProt: Q04759
Assay: In Vitro Inhibition Activity Assay: The compounds of formula I were tested for their activity on different PKC isoforms according to a published method (D. Geiges et al. Biochem. Pharmacol. 1997; 53:865-875) The assay is performed in a 96-well polypropylene microtiterplate (Costar 3794) that has been previously siliconized with Sigmacote (Sigma SL-2). The reaction mixture (50 μL) contains 10 μL of the relevant PKC isozyme together with 25 μL of the PKC inhibitor compound and 15 μL of a mix solution that contains 200 μg/mL protamine sulfate, 10 mM Mg(NO3)2, 10 μM ATP (Boehringer 519987) and 3750 Bq of 33P-ATP (Hartmann Analytic SFC301, 110 TBq/mmol) in 20 mM Tris-buffer pH 7.4+0.1% BSA. Incubation was performed for 15 minutes at 32° C. in a microtiterplate shaking incubator (Biolabo Scientific Instruments). Reaction was stopped by adding 10 μl of 0.5 M Na2EDTA, pH 7.4. 50 μl of mixture are pipetted onto a pre-wetted phosphocellulose paper (Whatmann 3698-915) under gentle pressure.
Measurement: IC50 = 0.36 nM (pChEMBL 9.4399995803833)
Descriptors: MW 442.53, ALogP 3.09, HBA 7, HBD 4, PSA 138.84, QED 0.38
Compound (SMILES): COc1cn(C(CC(C)C)C(=O)Nc2ccc(C(=O)O)cc2)c(=O)cc1-c1cc(Cl)ccc1OC(F)F
InChIKey: PONZAFSAZRCPMW-UHFFFAOYSA-N
Target: Plasma kallikrein (SINGLE PROTEIN), Homo sapiens
UniProt: P03952
Assay: Inhibition Assay: Test substances are dissolved in dimethyl sulphoxide and serially diluted in dimethyl sulphoxide (3000 μM to 0.0078 μM; resulting final concentrations in the test: 50 μM to 0.00013 μM). In each case 1 μl of the diluted substance solutions are placed into the wells of white microtitre plates from Greiner (384 wells). 20 μl of assay buffer (50 mM Tris/HCl pH 7.4; 100 mM sodium chloride solution; 5 mM of calcium chloride solution; 0.1% of bovine serum albumin) and 20 μl of plasma kallikrein from Kordia (0.6 nM in assay buffer) are then added successively. After 15 min of incubation, the enzyme reaction is started by addition of 20 μl of the substrate H-Pro-Phe-Arg-AMC dissolved in assay buffer (10 μl in assay buffer) from Bachem, the mixture is incubated at room temperature (22° C.) for 30 min and fluorescence is then measured (excitation: 360 nm, emission: 460 nm).
Measurement: IC50 = 360 nM (pChEMBL 6.440000057220459)
Descriptors: MW 534.94, ALogP 5.7, HBA 6, HBD 2, PSA 106.86, QED 0.35
Compound (SMILES): COc1ccc(C2(C(F)(F)F)CC(c3ccc(C)cc3)=C(C(=O)Nc3ccc(F)cc3)C(=O)N2)cc1
InChIKey: GKGZTRFCNASLKC-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: MGAT SPA Assay: MGAT2 enzyme was assayed using membranes isolated from Sf9 cells expressing the recombinant human MGAT2 cDNA with 2-monooleoylglycerol and [3H]-oleoyl-CoA as substrates as described by Seethala et al. [Anal. Biochem., 383(2):144-150 (Dec. 15, 2008)]. Briefly, the assays were conducted in 384-well plates in a total volume of 30 uL at 25° C. In each assay, 200 ng of recombinant human MGAT2 membrane was incubated with 10 uM of 2-monooleoylglycerol and 15 uM of [3H]-oleoyl-CoA in 100 mM potassium phosphate (pH 7.4) for 20 min with various concentrations of compounds delivered in DMSO. The assay was terminated by the addition of 20 ul of Stopping Solution (7.5 mg/ml Yttrium Oxide Polylysine beads, 3.3 mg/ml Fraction V BSA and 200 uM Mercuric chloride in 50 mM HEPES, pH 7.4). The signal was measured 1 h after quenching the reaction using LEADSEEKERSM for 5 minutes. To calculate the degree of inhibition, the zero level of enzyme activity (blank) was defined.
Measurement: IC50 = 170 nM (pChEMBL 6.769999980926514)
Descriptors: MW 498.48, ALogP 5.51, HBA 3, HBD 2, PSA 67.43, QED 0.36
Compound (SMILES): O=C(Nc1nncs1)N1CCC(c2nnn3cnc4[nH]ccc4c23)CC1
InChIKey: LBYCHMOAEQNCHG-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 64 nM (pChEMBL 7.190000057220459)
Descriptors: MW 369.41, ALogP 1.87, HBA 8, HBD 2, PSA 116.99, QED 0.56
Compound (SMILES): Nc1nc2cccc(Cl)c2c2nc(-c3ccco3)nn12
InChIKey: LTHNIAHTIWLCEM-UHFFFAOYSA-N
Target: Adenosine receptor A2a (SINGLE PROTEIN), Homo sapiens
UniProt: P29274
Assay: Radioligand Binding Assays: Radioligand binding assays at hA1, hA2A, and hA3ARs were performed according to the procedures described in Gao, Z. G., et al., Biochem Pharmacol. 2003; 65: 1675-84, using the radiolabeled agonists [3H]R-PIA from Moravek Biochemicals (Brea, Calif.) or [3H]CGS21680 and [125I]I-AB-MECA from PerkinElmer (Waltham, Mass.) for the hA1, hA2AAR, and hA3AR assays, respectively.
Measurement: Ki = 1.2 nM (pChEMBL 8.920000076293945)
Descriptors: MW 285.69, ALogP 2.77, HBA 6, HBD 1, PSA 82.24, QED 0.58
Compound (SMILES): CC(Nc1ncnc2[nH]cnc12)c1cc2cccc(Cl)c2c(=O)n1-c1ccccc1
InChIKey: SJVQHLPISAIATJ-UHFFFAOYSA-N
Target: PI3-kinase p110-delta/p85-alpha (PROTEIN COMPLEX), Homo sapiens
UniProt: O00329
Assay: Inhibition of N-terminal His6-tagged recombinant full-length human p110delta/untagged recombinant full length p85alpha expressed in baculovirus infected Sf21 cells using phosphatidylinositol as substrate in presence of gamma-32P-ATP after 5 mins by thin layer chromatographic method
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 416.87, ALogP 4.48, HBA 6, HBD 2, PSA 88.49, QED 0.45
Compound (SMILES): COCc1ccc(C)c(N(C)c2nc(Nc3cc(N4CCOCC4)cc(S(C)(=O)=O)c3)ncc2F)c1
InChIKey: BCGFTSYNPJUGDC-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase PAK 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13153
Assay: Inhibition of PAK1 (unknown origin) by ATP-kinaseGlo assay
Measurement: IC50 = 189 nM (pChEMBL 6.71999979019165)
Descriptors: MW 515.61, ALogP 3.82, HBA 9, HBD 1, PSA 96.89, QED 0.48
Compound (SMILES): COCCCCn1c(C(=O)N(CC(C)C)[C@@H]2CNC[C@H](C(=O)N3CCOCC3)C2)nc2ccccc21.Cl
InChIKey: PUXOYQIZZIWCHH-NSLUPJTDSA-N
Target: Renin (SINGLE PROTEIN), Homo sapiens
UniProt: P00797
Assay: Inhibition of renin in human plasma by radioimmunoassay
Measurement: IC50 = 2.1 nM (pChEMBL 8.680000305175781)
Descriptors: MW 499.66, ALogP 2.4, HBA 7, HBD 1, PSA 88.93, QED 0.51
Compound (SMILES): CN1C(=O)CCc2ccc(OCCCCN3CCN(c4cccc5sccc45)CC3)cc21
InChIKey: PUJLFURZMNJZQS-UHFFFAOYSA-N
Target: 5-hydroxytryptamine receptor 2A (SINGLE PROTEIN), Rattus norvegicus
UniProt: P14842
Assay: Serotonin 5-HT2A Receptor Binding Assay: The assay was performed according to the method by Leysen J E et al. (Leysen J E, Niemegeers C J E, Van Nueten J M and Laduron P M. [3H] Ketanserin (R 41 468), a selective 3H-ligand for serotonin 2 receptor binding sites. Mol. Pharmacol., 1982, 21: 301-314). The binding assay was performed using 40 ul of the membrane specimen, 20 ul of [3H]-Ketanserin (final concentration 1 to 3 nM), 20 ul of a test drug and 50 mM Tris-hydrochloric acid buffer (pH 7.4) so that the total amount was 200 ul (final dimethylsulfoxide concentration 1%). The reaction was performed at 37° C. for 20 minutes and terminated by conducting suction filtration with a cell harvester on a glass fiber filter plate.The filter plate made of glass fiber was washed with 50 mM Tris-hydrochloric acid buffer (pH 7.4), and after dried, a microplate liquid scintillation cocktail was added and the radioactivity was measured with a microplate scintillation counter.
Measurement: Ki = 8 nM (pChEMBL 8.100000381469727)
Descriptors: MW 449.62, ALogP 4.79, HBA 5, HBD 0, PSA 36.02, QED 0.49
Compound (SMILES): COc1cc(OC)cc(N(CC(N)=O)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: CRMQCPICYHHFIL-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 418.46, ALogP 2.67, HBA 8, HBD 1, PSA 108.39, QED 0.49
Compound (SMILES): CCN1CC(C)(C)Oc2nc(N3C4CCC3COC4)nc(-c3ccc(NC(=O)Nc4ccc(N5C[C@@H]6C[C@H]5CO6)nc4)c(F)c3)c2C1=O
InChIKey: YPTNSXVHACWVBJ-MXXWWZHRSA-N
Target: Phosphatidylinositol 3-kinase regulatory subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: P27986
Assay: Enzyme Assay: The test uses a lucifeurerin/luciferase system to measure the concentration of ATP and its consumption during the enzymatic reaction. The test is performed in 96-well format (Corning/Costar 96 black flat-bottomed half-wells plate, ref. 3694) in a total volume of 30 μl. To 1 μl of inhibitor in 100% DMSO are added (final concentrations) 50 μM of the substrate PIP2 ((L-α-phosphatidyl-D-myoinositol 4,5-bisphosphate, Echelon 117P-4516-0500), 2 μM of ATP and 1.7 μg/ml of PI3Kα, (p110α/p85α, Invitrogen PV4788) in a buffer of Tris/HCl 50 mM pH 7.5, EGTA 1 mM, MgCl2 10 mM, Chaps 0.03%, 1 mM DTT). After 90 minutes, the reaction is quenched by adding 20 μl/well of KinaseGlo reagent (Promega V6713). After 10 minutes in the dark, the luminescence is read on the PHERAStar microplate reader (reading at 0.8 sec/well).
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 658.74, ALogP 4.3, HBA 10, HBD 2, PSA 134.28, QED 0.4
Compound (SMILES): [C-]#[N+]c1cncc(-c2cc(Cl)c([C@]3(C)CS(=O)(=O)C4(CC4)C(=N)N3)s2)c1
InChIKey: QPDBFNARPOLNRB-INIZCTEOSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: HTRF FRET Assay: Inhibitor IC50s at purified human autoBACE-2 are determined in a time-resolved endpoint proteolysis assay that measures hydrolysis of the QSY7-EISEVNLDAEFC-Eu-amide FRET peptide substrate (BACE-HTRF assay). BACE-mediated hydrolysis of this peptide results in an increase in relative fluorescence (RFU) at 620 nm after excitation with 320 nm light Inhibitor compounds, prepared at 3x the desired final concentration in 1x BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO are pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1x BACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30° C. The assay is initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 uM for 4 uM for autoBACE-2) prepared in 1x BACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C.
Measurement: Ki < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 406.92, ALogP 3.76, HBA 5, HBD 2, PSA 87.27, QED 0.74
Compound (SMILES): Cc1ccccc1-n1c(Cn2nc(/C=C/C(N)=O)c3c(N)ncnc32)cc2cccc(C)c2c1=O
InChIKey: GXGMIHAYJQXAAS-ZHACJKMWSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P48736
Assay: Inhibition of recombinant human C-terminal His6-tagged p110gamma expressed in insect cells using phosphatidylinositol as substrate in presence of gamma-32P-ATP after 5 mins by thin layer chromatographic method
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 465.52, ALogP 2.88, HBA 8, HBD 2, PSA 134.71, QED 0.38
Compound (SMILES): COc1cc(-c2ccc(Cl)c(F)c2)c(F)cc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: ZCOUMXGRSDBUOU-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 = 8 nM (pChEMBL 8.100000381469727)
Descriptors: MW 543.94, ALogP 5.39, HBA 7, HBD 1, PSA 103.43, QED 0.31
Compound (SMILES): O=C(Nc1ccc(C(F)(F)F)cc1)N1CCC(F)(c2ncccc2Cl)CC1
InChIKey: USAYUWJTCYIKSX-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8NER1
Assay: pH-Based Assay: Two days prior to performing this assay, cells are seeded on poly-D-lysine-coated 96-well clear-bottom black plates (commercially available from Becton-Dickinson) at 75,000 cells/well in growth media containing 5 μM PonA (commercially available from Invitrogen) to induce expression of TRPV1. On the day of the assay, the plates are washed with 0.2 mL 1× Hank's Balanced Salt Solution (commercially available from Life Technologies) containing 1.6 mM CaCl2 and 20 mM HEPES, pH 7.4 ("wash buffer"), and loaded using 0.1 mL of wash buffer containing Fluo-4 (3 μM final concentration, commercially available from Molecular Probes). After 1 h, the cells are washed twice with 0.2 mL wash buffer and resuspended in 0.05 mL 1× Hank's Balanced Salt Solution (commercially available from Life Technologies) containing 3.5 mM CaCl2 and 10 mM Citrate, pH 7.4 ("assay buffer"). Plates are then transferred to a FLIPR for assay. The test compound is diluted in assay buffer, and 50 μL of the resultant solution is added to the cell plates and the solution is monitored for two minutes. The final concentration of the test compound is adjusted to range from about 50 picoM to about 3 μM. Agonist buffer (wash buffer titrated with 1N HCl to provide a solution having a pH of 5.5 when mixed 1:1 with assay buffer) (0.1 mL) is then added to each well, and the plates are incubated for 1 additional minute.
Measurement: IC50 = 51.2 nM (pChEMBL 7.289999961853027)
Descriptors: MW 401.79, ALogP 5.25, HBA 2, HBD 1, PSA 45.23, QED 0.7
Compound (SMILES): CC1CC2(CCC1CC(=O)O)CCN(c1cc(OC(F)(F)F)ccc1Cl)CC2
InChIKey: NLNNWVPBXBVTLL-UHFFFAOYSA-N
Target: Free fatty acid receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5NUL3
Assay: Agonist activity at human GPR120 short isoform expressed in CHOK1 cells assessed as induction of IP1 accumulation after 60 mins by HTRF assay
Measurement: EC50 = 4100 nM (pChEMBL 5.389999866485596)
Descriptors: MW 419.87, ALogP 5.74, HBA 3, HBD 1, PSA 49.77, QED 0.68
Compound (SMILES): CCCc1nn(C)c2c(=O)[nH]c(-c3cc(NC4CCN(c5ccc(C(=O)NO)cc5)CC4)ccc3OCC)nc12
InChIKey: WWAXWNFQIUYRFG-UHFFFAOYSA-N
Target: Histone deacetylase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13547
Assay: Inhibition of full length recombinant human C-terminal His/Flag-tagged HDAC1 expressed in baculovirus infected Sf9 cells using fluorogenic HDAC substrate after 30 mins by fluorescence analysis
Measurement: IC50 = 7810 nM (pChEMBL 5.110000133514404)
Descriptors: MW 545.64, ALogP 3.87, HBA 9, HBD 4, PSA 137.4, QED 0.18
Compound (SMILES): CC(=O)Nc1ccc(Nc2nccc(Nc3ccccc3Cl)n2)cc1
InChIKey: OXRRJTBMYSVLSJ-UHFFFAOYSA-N
Target: Aurora kinase A (SINGLE PROTEIN), Homo sapiens
UniProt: O14965
Assay: In Vivo Aurora A inhibitory Assay ( MDA-MB-468): Specifically, MDA-MB-468 cells (American Type Culture Collection) were maintained in Dulbecco's modified Eagle's medium (DMEM) plus 10% fetus bovine serum (FBS) (Invitrogen, US) at 37° C., 5% CO2. The cells were plated in 6 cm dishes at a density of 2x105 cells/dish. Cells were then treated with the compounds (0-10 uM); DMSO was used as a negative control and 0.5 uM VX-680 (Tozasertib) (Selleck Chemical LLC) was used as a positive control. Cells were harvested after 2 h of treatment and processed for SDS-PAGE and western blotting as described previously (Gizatullin et al., "The Aurora kinase inhibitor VX-680 induces endoreduplication and apoptosis preferentially in cells with compromised p53-dependent postmitotic checkpoint function" Cancer Res. (2006) 66 (15):7668-77). After electro-transfer onto nitrocellulose membranes, the membranes were blocked at room temperature for 1 h with TBS containing 5% (w/v) milk and then washed with a mixture of TBS containing 0.2% Tween 20 (Sigma). The membranes were then gently shaken at 4° C. overnight with anti-phospho histone H3 (Ser 10) antibody (9701, Cell Signaling), and anti-GAPDH monoclonal antibody (E10086CF, Covance) diluted in TBS containing 5% BSA. The membranes were then incubated with HRP conjugated anti-rabbit or anti-mouse IgG antibody (Jackson ImmunoResearch Lab.) at room temperature for 1 h followed by washing with Tween 20-PBS. The membranes were washed again with PBS and developed with the ECL system (PerkinElmer) as described previously (Berndt et al., "The Akt activation inhibitor TCN-P inhibits Akt phosphorylation by binding to the PH domain of Akt and blocking its recruitment to the plasma membrane." Cell Death Differ. (2010) 17(11):1795-804).
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 353.81, ALogP 4.58, HBA 5, HBD 3, PSA 78.94, QED 0.63
Compound (SMILES): CC(C)n1cc(-c2cnc3[nH]cc(-c4cc(C(C)(O)c5ccccc5)nc(N)n4)c3c2)cn1
InChIKey: JDBDJPSCRBWYRW-UHFFFAOYSA-N
Target: 3-phosphoinositide-dependent protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O15530
Assay: Luminex Assay: The cellular assay for the determination of the PDK1 kinase activity is carried out as a Luminex assay in the 96-well format. PC3 cells are sown at 20,000 cells per well in 100 ul of medium (45% of RPMI1460/45% of Ham's F12/10% of FCS) and incubated on the following day for 30 min with a serial dilution of the test substance (7 concentrations) under serum-free conditions. The cells are subsequently lysed using 90 ul of lysis buffer (20 mM tris/HCl pH 8.0, 150 mM NaCl, 1% of NP40, 10% of glycerol, 1% of phosphatase inhibitor I, 1% of phosphatase inhibitor II, 0.1% of protease inhibitor cocktail III, 0.01% of benzonase) per well, and the lysates are separated off from insoluble cell constituents by means of centrifugation through a 96-well filter plate (0.65 um). The lysates are incubated overnight at 4° C. with shaking with Luminex beads to which an anti-total PKB antibody is coupled. The detection is carried out on the following day by addition of a phospho-T308-PKB antibody and a species-specific peroxidase-labelled secondary antibody.
Measurement: IC50 = 492 nM (pChEMBL 6.309999942779541)
Descriptors: MW 439.52, ALogP 4.3, HBA 7, HBD 3, PSA 118.53, QED 0.38
Compound (SMILES): CC(C(=O)Nc1ccc(-c2cc(=O)[nH][nH]2)cc1)n1cc(Cl)c(-c2cc(Cl)ccc2C#N)cc1=O
InChIKey: IVNAHVJAOFMEMQ-UHFFFAOYSA-N
Target: Plasma kallikrein (SINGLE PROTEIN), Homo sapiens
UniProt: P03952
Assay: Inhibition Assay: Test substances are dissolved in dimethyl sulphoxide and serially diluted in dimethyl sulphoxide (3000 μM to 0.0078 μM; resulting final concentrations in the test: 50 μM to 0.00013 μM). In each case 1 μl of the diluted substance solutions are placed into the wells of white microtitre plates from Greiner (384 wells). 20 μl of assay buffer (50 mM Tris/HCl pH 7.4; 100 mM sodium chloride solution; 5 mM of calcium chloride solution; 0.1% of bovine serum albumin) and 20 μl of plasma kallikrein from Kordia (0.6 nM in assay buffer) are then added successively. After 15 min of incubation, the enzyme reaction is started by addition of 20 μl of the substrate H-Pro-Phe-Arg-AMC dissolved in assay buffer (10 μl in assay buffer) from Bachem, the mixture is incubated at room temperature (22° C.) for 30 min and fluorescence is then measured (excitation: 360 nm, emission: 460 nm).
Measurement: IC50 = 110 nM (pChEMBL 6.960000038146973)
Descriptors: MW 494.34, ALogP 4.58, HBA 5, HBD 3, PSA 123.54, QED 0.38
Compound (SMILES): COc1cc(C(=O)NCc2ccc(F)cc2Cl)ccc1-c1cn[nH]c1
InChIKey: ICHFSXPUXZFBPH-UHFFFAOYSA-N
Target: Rho-associated protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O75116
Assay: In Vitro Assay: The effectiveness of compounds of the present invention as ROCK inhibitors can be determined in a 30 μL assay containing 20 mM HEPES, pH 7.5, 20 mM MgCl2, 0.015% Brij-35, 4 mM DTT, 5 μM ATP and 1.5 μM peptide substrate (FITC-AHA-AKRRRLSSLRA-OH). Compounds were dissolved in DMSO so that the final concentration of DMSO was <2%, and the reaction was initiated with Rho kinase variants. After incubation, the reaction was terminated by the addition of EDTA and the phosphorylated and non-phosphorylated peptides separated using a LABCHIP® 3000 Reader (Caliper Life Sciences). Controls consisted of assays that did not contain compound, and backgrounds consisted of assays that contained enzyme and substrate but had EDTA from the beginning of the reaction to inhibit kinase activity.
Measurement: IC50 = 2.54 nM (pChEMBL 8.600000381469727)
Descriptors: MW 359.79, ALogP 3.81, HBA 3, HBD 2, PSA 67.01, QED 0.73
Compound (SMILES): CN1Cc2c(Cl)cc(Cl)cc2[C@H](c2cccc(S(=O)(=O)NCCOCCOCCOCCNC(=O)CN3CCN(c4nc(N5CCN(CC(=O)NCCOCCOCCOCCNS(=O)(=O)c6cccc([C@@H]7CN(C)Cc8c(Cl)cc(Cl)cc87)c6)CC5)nc(N5CCN(CC(=O)NCCOCCOCCOCCNS(=O)(=O)c6cccc([C@@H]7CN(C)Cc8c(Cl)cc(Cl)cc87)c6)CC5)n4)CC3)c2)C1
InChIKey: JZIVLRCVVBXJIZ-DJYPZEJZSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P48764
Assay: Inhibition of human NHE3-mediated sodium-dependant hydrogen ion antiport in Opossum kidney cells preincubated in NH4Cl-HEPES buffer containing BCECF-AM dye followed by washout with ammonium free HEPES and subsequent addition of compound in presence of NHE1 antagonist EIPA by fluorescence assay
Measurement: IC50 = 199.53 nM (pChEMBL 6.699999809265137)
Descriptors: MW 2093.06
Compound (SMILES): CC(C)(C)Nc1nnc(-c2ccc3[nH]cc(-c4nccc(C5CC5)n4)c3c2)o1
InChIKey: BQKBRLMFCCUJQA-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P1W9
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 = 2.395 nM (pChEMBL 8.619999885559082)
Descriptors: MW 374.45, ALogP 4.76, HBA 6, HBD 2, PSA 92.52, QED 0.53
Compound (SMILES): CCC(=O)N1CCC(CC#N)(n2cc(C(N)=O)c(Nc3ccccc3)n2)CC1
InChIKey: JADBGAAVTXNBPR-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: HTRF Biochemical Assay: The reactions employed a common peptide substrate, LCB-EQEDEPEGDYFEWLW-NH2 (in-house). The basic assay protocol is as follows: First, 250 nL of diluted compounds in DMSO were dispensed into the wells of a dry 384-well Black plate (Greiner #781076) using a Labcyte Echo 555 acoustic dispenser. Subsequent reagent additions employed an Agilent Bravo. Next, 18 uL of 1.11x enzyme and 1.11x substrate in 1x assay buffer (Invitrogen kinase buffer # PV3189, 2 mM DTT, 0.05% BSA) were added to the wells and shaken and then preincubated for 30 minutes at room temperature to allow compound binding to equilibrate. After equilibration, 2 uL of 10xATP in 1x assay buffer was added to initiate the kinase reaction and the plates were shaken and then incubated at room temperature for 120 minutes. At the end of the incubation, 20 uL of 2x stop buffer (streptavidin-Dylight 650 (Thermo #84547B/100 mL), Eu-tagged pY20 antibody (Perkin Elmer #AD0067), EDTA, HEPES, and Triton) was added to quench the reaction.
Measurement: IC50 = 26 nM (pChEMBL 7.579999923706055)
Descriptors: MW 380.45, ALogP 2.37, HBA 6, HBD 2, PSA 117.04, QED 0.8
Compound (SMILES): NS(=O)(=O)c1ccc(O[C@H]2c3ccccc3C[C@@H]2N2CCCCC2)cc1
InChIKey: KPOFRWYZAASAIZ-PMACEKPBSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P48764
Assay: Inhibition of human NHE3-mediated sodium-dependant hydrogen ion antiport in Opossum kidney cells preincubated in NH4Cl-HEPES buffer containing BCECF-AM dye followed by washout with ammonium free HEPES and subsequent addition of compound in presence of NHE1 antagonist EIPA by fluorescence assay
Measurement: IC50 = 2511.89 nM (pChEMBL 5.599999904632568)
Descriptors: MW 372.49, ALogP 2.86, HBA 4, HBD 1, PSA 72.63, QED 0.9
Compound (SMILES): COc1cc(C(=O)N[C@@H](C)c2cccc3ccccc23)ccc1-c1cn[nH]c1
InChIKey: OEYSQZKVMVAGLO-HNNXBMFYSA-N
Target: Rho-associated protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O75116
Assay: In Vitro Assay: The effectiveness of compounds of the present invention as ROCK inhibitors can be determined in a 30 μL assay containing 20 mM HEPES, pH 7.5, 20 mM MgCl2, 0.015% Brij-35, 4 mM DTT, 5 μM ATP and 1.5 μM peptide substrate (FITC-AHA-AKRRRLSSLRA-OH). Compounds were dissolved in DMSO so that the final concentration of DMSO was <2%, and the reaction was initiated with Rho kinase variants. After incubation, the reaction was terminated by the addition of EDTA and the phosphorylated and non-phosphorylated peptides separated using a LABCHIP® 3000 Reader (Caliper Life Sciences). Controls consisted of assays that did not contain compound, and backgrounds consisted of assays that contained enzyme and substrate but had EDTA from the beginning of the reaction to inhibit kinase activity.
Measurement: IC50 = 2.32 nM (pChEMBL 8.630000114440918)
Descriptors: MW 371.44, ALogP 4.73, HBA 3, HBD 2, PSA 67.01, QED 0.53
Compound (SMILES): CCn1nc(-c2cccc(-c3cccc(O)c3)c2)cc(NC(=O)Nc2c(Cl)cncc2Cl)c1=O
InChIKey: DYSBLMVPFHWRLU-UHFFFAOYSA-N
Target: Adenosine receptor A2a (SINGLE PROTEIN), Homo sapiens
UniProt: P29274
Assay: Antagonist activity at adenosine A2a receptor (unknown origin)
Measurement: IC50 = 168 nM (pChEMBL 6.78000020980835)
Descriptors: MW 496.35, ALogP 5.65, HBA 6, HBD 3, PSA 109.14, QED 0.33