lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CN(C)Cc1ccc(-c2c(-c3cccc(OC(F)(F)F)c3)cc3n2[C@@H](CCN)CNC3=O)cc1
InChIKey: POOGYHHHUWNODC-IBGZPJMESA-N
Target: Serine/threonine-protein kinase pim-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P1W9
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 Supernatant was subsequently withdrawn and transferred into a counting plate, then evaluated by beta-counting.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 PIM2 assay was composed of HEPES 50 mM, at pH 7.5, with 1 mM MgCl2, 1 mM DTT, 3 uM Na3VO4, and 0.2 mg/mL BSA.Â  Full-length human PIM2 was expressed and purified as described in Fedorov O, et al., PNAS 2007 104, 51, 20523-28.
Measurement: IC50 = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 472.51, ALogP 4.42, HBA 5, HBD 2, PSA 72.52, QED 0.53
Compound (SMILES): CCC(CC)(NC(=O)c1ccc(C(F)(F)F)c(OCC2CCOCC2)n1)C(=O)NC
InChIKey: GGTIETYXENLZJP-UHFFFAOYSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1.times.HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37° C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30.degree. C. for 30 min Compounds were added to a final assay volume of 100 ul and incubated for 30 min at 30° C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 ul lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN.sub.3) and 50 ul detection solutions (20 uM mAb Alexa700-cAMP 1:1, and 48 uM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 431.46, ALogP 2.94, HBA 5, HBD 2, PSA 89.55, QED 0.66
Compound (SMILES): CCCN(CCC)C(=O)C1=Cc2ccc(C(=O)Nc3cncc(C#CCN)c3)cc2N=C(N)C1
InChIKey: XMXCMGODAVHNIF-UHFFFAOYSA-N
Target: Toll-like receptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NYK1
Assay: Pharmacological Assay: For TLR8 and TLR7 activity testing, HEK-Blue human TLR8 or TLR7 cells (Invivogen, San Diego, Calif., USA) are used, respectively. These cells are designed for studying the stimulation of human TLR8 or TLR7 by monitoring the activation of NF-κB. A SEAP (secreted embryonic alkaline phosphatase) reporter gene is placed under the control of the IFN-b minimal promoter fused to five NF-κB and AP-1-binding sites. Therefore the reporter expression is regulated by the NF-κB promoter upon stimulation of human TLR8 or TLR7 for 20 hours. The cell culture supernatant SEAP reporter activity was determined using Quanti Blue kit (Invivogen, San Diego, Ca, USA) at a wavelength of 640 nm, a detection medium that turns purple/blue in the presence of alkaline phosphatase.
Measurement: EC50 > 100000 nM (pChEMBL 4)
Descriptors: MW 458.57, ALogP 3.07, HBA 6, HBD 3, PSA 126.7, QED 0.55
Compound (SMILES): Cc1c(C(=O)Nc2cccc(C(F)(F)F)c2)cc(C(C)(C)C)n1C1CCN(CCO)CC1
InChIKey: IUEFZOBSIYWIPD-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9HBA0
Assay: Inhibition Assay: Test Example 1: (1) The TRPV4 inhibitory activity was measured using CHO-K1 cells stably expressing human TRPV4 (hTRPV4/CHO cells). (2) In the day before the experimental day, frozen cell were thawed and washed with the culture medium (MEM-α, 10% FBS, 2 mmol/L GlutaMax, 50 unit Penicillin, 50 μg/mL Streptomycin). Then cells were suspended in the culture medium. (3) The 384-well plates which hTRPV4/CHO cells were seeded at densities of 4000 cells/well in the culture medium and cultured in a CO2 incubator in the presence of 5% CO2 at 37° C. for overnight was used as assay plates. (4) The assay plate was washed with assay buffer (Hanks, 20 mmol/L HEPES, 2.5 mmol/L probenecid, pH7.4), and the buffer were remained at 20 μL/well. (5) 10 μl of dye loading buffer (9 μmol/L Fluo 4-AM, 0.09% Pluronic F-127/assay buffer) was added to each well, then the assay plate was incubated in a CO2 incubator in the presence of 5% CO2 at 37° C. for one hour. (final 3 μmol/L Fluo4-AM). (6) The assay plate was washed with the assay buffer, and the buffers were remained at 20 μL/well. Then the assay plate was incubated for 10 min at room temperature. (7) Diluted compound solution was dispensed to each well in the compound plate, and mixed with built-in Pipette and Mixer of the Fluorescence analysis system FLIPR TETRA (Molecular devices). (8) After incubation for 5 min, 20 μL of 4α-PDD solution were applied to each well of assay plate and mixed with the FLIPR TETRA. (final 1 Î¼mol/L 4α-PDD). (9) The fluorescent intensity was measured with FLIPR TETRA system for 10 min from the point of time addition the compound solution, at Ex 470-495 nm, Em 515-575 nm wavelength.
Measurement: IC50 = 6600 nM (pChEMBL 5.179999828338623)
Descriptors: MW 451.53, ALogP 4.99, HBA 4, HBD 2, PSA 57.5, QED 0.67
Compound (SMILES): CC(C)(O)c1cn(-c2ccc(-c3cc(F)c(CO)c(S(C)(=O)=O)c3)cc2)c(C(C)(C)c2c(Cl)cccc2Cl)n1
InChIKey: LODWYHGIAQYCAL-UHFFFAOYSA-N
Target: Retinoic acid receptor RXR-alpha/oxysterols receptor LXR-alpha (PROTEIN COMPLEX), Homo sapiens
UniProt: P19793
Assay: Displacement of [3H]-24,25-epoxycholesterol from human LXRalpha/RXRalpha expressed in baculovirus infected Sf9 cells by scintillation proximity analysis
Measurement: Ki = 38 nM (pChEMBL 7.420000076293945)
Descriptors: MW 591.53, ALogP 6.43, HBA 6, HBD 2, PSA 92.42, QED 0.26
Compound (SMILES): CN[C@@H](C)C(=O)N[C@@H]1C(=O)N(Cc2c(C3CC3)cnc3ccccc23)c2ccccc2N(C(=O)c2ccc(C(C)=O)cc2)[C@H]1C
InChIKey: FSSFEVHWWBPTSY-RACYCIBVSA-N
Target: E3 ubiquitin-protein ligase XIAP (SINGLE PROTEIN), Homo sapiens
UniProt: P98170
Assay: TR-FRET Assay: The peptide AVPIAQKSEK-(ε-biotin)-OH 1:2 TFA ("Peptide A") was identified as a substrate for the TR-FRET assay by screening the 6× Histidine-tagged BIR2 domain and BIR3 domain of XIAP against a set of 29 peptides synthesized based on sequences reported by Sweeny et al. (Biochemistry, 2006, 45, 14740 14748). The peptides were labeled with the fluorescent tags FITC or TAMRA and Kd values were determined by fluorescence polarization assay. The sequence AVPIAQKSEK was identified as optimal for using in an assay. The peptide sequence was derivatized with biotin to provide AVPIAQKSEK-(ε-biotin)-OH 1:2 TFA as the substrate for the TR-FRET assay. Ten nanomolar of 6× Histidine-tagged BIR2 domain, corresponding to amino acids 124-240 of XIAP, or BIR3 domain, corresponding to amino acids 241-356 of XIAP, was mixed with 20 nM of the peptide AVPIAQKSEK-(ε-biotin)-OH 1:2 TFA, in the presence of 50 mM Tris-C1, pH 7.5, 100 mM NaCl, 1 mM dithiothreitol (DTT) and 0.1 mg/mL bovine serum albumin (BSA). Following a 45 mM incubation at 37° C., Europium-Streptavidin and Allophycocyanin conjugated anti-Histidine antibody were added to a final concentration of 1.5 nM and 15 nM, respectively. Time-resolved fluorescence resonance energy transfer (TR-FRET) signals were measured 1 hour later at room temperature. Test compound potency was assessed at 10 serially diluted concentrations.
Measurement: IC50 = 15100 nM (pChEMBL 4.820000171661377)
Descriptors: MW 603.72, ALogP 4.99, HBA 6, HBD 2, PSA 111.71, QED 0.27
Compound (SMILES): O=C(O)[C@@H]1CCN1c1nc2ccccc2n([C@H]2C[C@H]3CC[C@@H](C2)N3[C@@H]2C[C@@H]3CCCC[C@@H](C3)C2)c1=O
InChIKey: CUUNNNJSQSWPEX-BIPLSNKXSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: In Vitro Mu-Opioid Receptor Binding Assay: Radioligand binding assays were conducted using freshly thawed membranes expressing human μ-receptors (Perkin Elmer, Shelton, Conn.). Radioligand dose-displacement binding assays for human μ-opioid receptors used 0.2 nM [3H]-diprenorphine (NEN, Boston, Mass.), with 5-20 mg membrane protein/well in a final volume of 500 μL binding buffer (10 mM MgCl2, 1 mM EDTA, 5% DMSO, 50 mM HEPES, pH 7.4). Reactions were carried out in the absence or presence of increasing concentrations of unlabeled naloxone. All reactions were conducted in 96-deep well polypropylene plates for 1-2 hr at about 25 °C. Binding reactions were terminated by rapid filtration onto 96-well Unifilter GF/C filter plates (Packard, Meriden, Conn.) presoaked in 0.5% polyethylenimine using a 96-well tissue harvester (Brandel, Gaithersburg, Md.) followed by performing three filtration washes with 500 μL of ice-cold binding buffer. Filter plates were subsequently dried at 50 °C. for 2-3 hours. BetaScint scintillation cocktail (Wallac, Turku, Finland) was added (50 μL/well), and plates were counted using a Packard Top-Count for 1 min/well.
Measurement: Ki = 259 nM (pChEMBL 6.590000152587891)
Descriptors: MW 490.65, ALogP 4.59, HBA 6, HBD 1, PSA 78.67, QED 0.68
Compound (SMILES): Cc1cc(C)c(N(C)c2nc(Nc3ccc(N4CCN(C)CC4)cc3)ncc2F)cc1CO
InChIKey: MUTUMOBIBVSADG-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 = 33 nM (pChEMBL 7.480000019073486)
Descriptors: MW 450.56, ALogP 3.99, HBA 7, HBD 2, PSA 67.76, QED 0.59
Compound (SMILES): O=C(OC1C2CC3CC(C2)CC1C3)N1CCC(CO)(c2ccccc2)CC1
InChIKey: WEJDNNCKEWKRDO-UHFFFAOYSA-N
Target: 11-beta-hydroxysteroid dehydrogenase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P28845
Assay: Scintillation Proximity Assay: Inhibition of microsomal preparations of 11.beta.-HSDl by compounds of the present invention, as described essentially previously (K. Solly, SS Mundt, HJ Zokian, GJ Ding, A. Hermanowski-Vosatka, B. Strulovici, and W. Zheng, High-Throughput Screening of 11-Beta-Hydroxyseroid Dehydrogenase Type 1 in Scintillation Proximity Assay Format. Assay Drug Dev Technol 3 (2005) 377-384) was measured. All reactions were carried out in a clear flexible PET Microbeta plates 96-well at room temperature (PerkinElmer). Assay, substrate solution (50mM HEPES, pH7.4,100mM KCl, 5mM NaCl, 2mM MgCl 2, 2mM NADPH and 160nM [3 H] cortisone (1Ci / mmol)) a 49ul and distribution, and starting from the 0.1mM on to start by combining the semi-log (half-log) multiple of the incremental test compound 1uL in DMSO previously diluted (8 points). After pre-incubation of 10 minutes, incubated human 11-HSD1 overexpress the enzyme containing CHO cells isolated from microsomes solution 50uL.
Measurement: IC50 = 4.44 nM (pChEMBL 8.350000381469727)
Descriptors: MW 369.51, ALogP 3.97, HBA 3, HBD 1, PSA 49.77, QED 0.88
Compound (SMILES): Fc1ccc(C(SCCN2CCN(Cc3ccccc3)CC2)c2ccc(F)cc2)cc1
InChIKey: ATOFTVWSYSKDIM-UHFFFAOYSA-N
Target: Sodium-dependent serotonin transporter (SINGLE PROTEIN), Rattus norvegicus
UniProt: P31652
Assay: Displacement of [3H]citalopram from SERT in Sprague-Dawley rat brain stem membranes incubated for 60 mins by radioligand binding assay
Measurement: Ki = 949 nM (pChEMBL 6.019999980926514)
Descriptors: MW 438.59, ALogP 5.61, HBA 3, HBD 0, PSA 6.48, QED 0.45
Compound (SMILES): NC(=O)CC1(NC(=O)c2cnc(N3CC(F)(F)C3)c(OCC3CC3)n2)COC1
InChIKey: PSJUKZIHVLIJAP-UHFFFAOYSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1×HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37° C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30° C. for 30 min. Compounds were added to a final assay volume of 100 μl and incubated for 30 min at 30° C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 μl lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN3) and 50 μl detection solutions (20 μM mAb Alexa700-cAMP 1:1, and 48 μM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 397.38, ALogP 0.09, HBA 7, HBD 2, PSA 119.67, QED 0.64
Compound (SMILES): CC(C)OC(=O)CCC(=O)Nc1cccc(-c2n[nH]c3ccc(NS(=O)(=O)c4ccccc4)cc23)c1
InChIKey: QSKHBRBUJVMKMR-UHFFFAOYSA-N
Target: Aurora kinase B (SINGLE PROTEIN), Homo sapiens
UniProt: Q96GD4
Assay: Inhibition of human recombinant full-length His-tagged Aurora B expressed in baculovirus expression system
Measurement: IC50 = 220 nM (pChEMBL 6.659999847412109)
Descriptors: MW 506.58, ALogP 4.7, HBA 6, HBD 3, PSA 130.25, QED 0.28
Compound (SMILES): CCN(C(=O)c1cnc(C(C)C)nc1NCc1ccco1)C1CCCNC1.Cl.Cl
InChIKey: LZWYNBQNIPHINN-UHFFFAOYSA-N
Target: Renin (SINGLE PROTEIN), Homo sapiens
UniProt: P00797
Assay: Inhibition of recombinant human renin expressed in FreeStyle 293 expression system using FITC-epsilonAcp-Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu-Val-Ile-His-Asn-Arg-NH2 as substrate preincubated for 15 mins followed by substrate addition measured after 30 mins by microchip-type capillary electrophoresis method
Measurement: IC50 = 470 nM (pChEMBL 6.329999923706055)
Descriptors: MW 371.49, ALogP 3.02, HBA 6, HBD 2, PSA 83.29, QED 0.78
Compound (SMILES): CC(=O)c1ccccc1NC(=O)/C=C\c1ccccc1
InChIKey: MJBHMIDHYIBRTA-QXMHVHEDSA-N
Target: Shiga toxin subunit A (SINGLE PROTEIN), Shigella dysenteriae
UniProt: Q9FBI2
Assay: Inhibition of Shigella dysenteriae type 1 Shiga toxin A subunit in African green monkey Vero cells assessed as inhibition of Stx-induced cytotoxicity pre-treated with compound for 1 hr followed by Stx exposure for 24 hrs by neutral red uptake assay
Measurement: IC50 = 2630 nM (pChEMBL 5.579999923706055)
Descriptors: MW 265.31, ALogP 3.54, HBA 2, HBD 1, PSA 46.17, QED 0.68
Compound (SMILES): CC1(Nc2ncc(C(F)(F)F)cn2)CCCC1NC(=O)c1ncccc1-c1ncccn1
InChIKey: UGCBWTSZNUXSMP-UHFFFAOYSA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: Fluorescent Imaging Plate Reader (FLIPR) Assay: Orexin antagonist activity was determined by measuring changes in intracellular calcium levels using a Ca2+ sensitive fluorescent dye. The changes in fluorescent signal were monitored by Fluorescent Imaging Plate Reader (FLIPR®) technology available from Molecular Devices, LLC, U.S.A. Orexin mediated increases in intracellular Ca2+ concentration were readily detected upon activation with orexin-A. Twenty-four hours prior to the assay, RBL-2H3 cells stably expressing either human orexin receptor 1 or human orexin receptor 2 were seeded in cell culture medium in black, clear-bottom 384-well plates (commercially available from Corning Inc., U.S.A.) and grown overnight at 37 °C., 5% CO2. On the day of the assay, cell culture media was removed and cells were loaded with Calcium 5 Dye (commercially sold by Molecular Devices, LLC, U.S.A.) for 1 hour at 37 °C., 5% CO2.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 443.43, ALogP 3.5, HBA 7, HBD 2, PSA 105.58, QED 0.62
Compound (SMILES): CNCCN(c1cc(OC)cc(OC)c1)c1ccc2ncc(-c3cnn(CC(=O)NC)c3)nc2c1
InChIKey: YIKYPGSFLXUTQT-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 475.55, ALogP 2.61, HBA 9, HBD 2, PSA 106.43, QED 0.36
Compound (SMILES): O=C(O)c1ccc(COc2ccc(C(=O)Nc3ccc(C4CNCCO4)cc3)cc2)cc1
InChIKey: RSUNWYDYKLNWCM-UHFFFAOYSA-N
Target: Trace amine-associated receptor 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q923Y9
Assay: Radioligand Binding Assay: HEK-293 cells stably expressing rat TAAR1 were maintained at 37° C. and 5% CO2 in DMEM high glucose medium, containing fetal calf serum (10%, heat inactivated for 30 min at 56° C.), penicillin/streptomycin (1%), and 375 ug/ml geneticin (Gibco). Cells were released from culture flasks using trypsin/EDTA, harvested, washed twice with ice-cold PBS (without Ca2+ and Mg2+), pelleted at 1,000 rpm for 5 min at 4° C., frozen and stored at -80° C. Frozen pellets were suspended in 20 ml HEPES-NaOH (20 mM, pH 7.4) containing 10 mM EDTA and homogenized with a Polytron (PT 6000, Kinematica) at 14,000 rpm for 20 s. The homogenate was centrifuged at 48,000Ã¿g for 30 min at 4° C. Subsequently, the supernatant was removed and discarded, and the pellet resuspended in 20 ml HEPES-NaOH (20 mM, pH 7.4) containing 0.1 mM EDTA using the Polytron (20 s at 14,000 rpm). This procedure was repeated and the final pellet resuspended in HEPES-NaOH containing 0.1 mM EDTA and homogenized using the Polytron. Typically, aliquots of 2 ml membrane portions were stored at -80° C. With each new membrane batch the dissociation constant (Kd) was determined via a saturation curve. The TAAR1 radioligand 3[H]â¿¿(S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine (described in WO 2008/098857) was used at a concentration equal to the calculated Kd value, that was usually around 2.3 nM, resulting in the binding of approximately 0.2% of the radioligand and a specific binding representing approximately 85% of the total binding. Nonspecific binding was defined as the amount of 3[H]â¿¿(S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine bound in the presence of 10 uM unlabeled ligand. All compounds were tested at a broad range of concentrations (10 pM to 10 uM) in duplicates. The test compounds (20 ul/well) were transferred into a 96 deep well plate (TreffLab), and 180 ul of HEPES-NaOH (20 mM, pH 7.4) containing MgCl2 (10 mM) and CaCl2 (2 mM) (binding buffer), 300 ul of the radioligand 3[H]â¿¿(S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine at a concentration of 3.3xKd in nM and 500 ul of the membranes (resuspended at 50 ug protein per ml) added. The 96 deep well plates were incubated for 1 hr at 4° C. Incubations were terminated by rapid filtration through Unifilter-96 plates (Packard Instrument Company) and glass filters GF/C (Perkin Elmer) presoaked for 1 hr in polyethylenimine (0.3%) and washed 3 times with 1 ml of cold binding buffer. After addition of 45 ul of Microscint 40 (PerkinElmer) the Unifilter-96 plate was sealed and after 1 hr the ratioactivity counted using a TopCount Microplate Scintillation Counter (Packard Instrument Company).
Measurement: Ki = 295.8 nM (pChEMBL 6.53000020980835)
Descriptors: MW 432.48, ALogP 3.88, HBA 5, HBD 3, PSA 96.89, QED 0.52
Compound (SMILES): C[C@@]12[C@@H]3CC[C@@H](C3=O)[C@@H]1CN(c1ccc(C#N)c(C(F)(F)F)c1)S2(=O)=O
InChIKey: LGYLNMVCFJRFAT-UNIGVISCSA-N
Target: Androgen receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P10275
Assay: Displacement of [3H]DHT from androgen receptor in human MDA-MB-453 cells
Measurement: Ki = 8 nM (pChEMBL 8.100000381469727)
Descriptors: MW 384.38, ALogP 2.71, HBA 4, HBD 0, PSA 78.24, QED 0.75
Compound (SMILES): N#CCC1(n2cc(C(N)=O)c(Nc3ccc(F)cc3)n2)CCN(Cc2ccc(F)cn2)CC1
InChIKey: DYUKTLRGURYWGF-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 = 73 nM (pChEMBL 7.139999866485596)
Descriptors: MW 451.48, ALogP 3.3, HBA 7, HBD 2, PSA 112.86, QED 0.57
Compound (SMILES): COc1cc2ccccc2cc1CCc1c(C(=O)O)[nH]c(=O)[nH]c1=O.NCCO
InChIKey: GLLSWLQKNYNUMZ-UHFFFAOYSA-N
Target: Dihydroorotate dehydrogenase (quinone), mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: Q02127
Assay: Inhibition of human dihydroorotate dehydrogenase assessed as reduction in orotate production using L-dihydroorotate as substrate measured after 20 mins by spectrophotometric method
Measurement: IC50 = 39.8 nM (pChEMBL 7.400000095367432)
Descriptors: MW 340.34, ALogP 1.71, HBA 4, HBD 3, PSA 112.25, QED 0.65
Compound (SMILES): NC(N)=Nc1nc(-c2ccc(N3CCN(CCOCCOCCOCCNC(=O)NCCOCCOCCNC(=O)CN(CCN(CC(=O)O)CC(=O)NCCOCCOCCNC(=O)NCCOCCOCCOCCN4CCN(c5ccc(-c6nc(N=C(N)N)nc7ccc(Cl)cc67)cc5)CC4)CC(=O)O)CC3)cc2)c2cc(Cl)ccc2n1
InChIKey: CAWRFGLWEYEAGM-UHFFFAOYSA-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 = 12.59 nM (pChEMBL 7.900000095367432)
Descriptors: MW 1718.81
Compound (SMILES): CNC(=O)c1cccc(Nc2ncnc(N[C@H](C)CCc3ccccc3)n2)c1
InChIKey: UEDLMLQFOLKQKR-OAHLLOKOSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Cell-Based Assay: Compounds were tested on human Nav1.7 (hNav1.7) expressed stably in 293 cells, which have no endogenous sodium channels. Two forms of whole-cell patch-clamp assays were used. In both, peak current through Nav1.7 was evoked by a depolarizing voltage pulse, and currents were monitored as a function of time in response to exposure to several different concentrations of drug. Automated electrophysiology uses the PatchXpress 7000A system. Up to sixteen cells at a time were voltage-clamped in parallel, and the steady-state voltage dependence of inactivation was determined experimentally for each cell. For each cell, fractional inactivation was set at 20%, and peak current through Nav1.7 as a function of time was measured in response to increasing concentrations of test compound. Manual electrophysiology refers to conventional whole-cell patch-clamping (Hamill et al., 1981). After formation of the whole-cell clamp, cells were lifted off the bottom of the dish with the patch pipette and positioned directly in front of an array of glass microperfusion tubes each with internal diameter about 250 microns, with individual compound concentrations flowing from each tube. Current as a function of time in response to increasing drug concentrations was measured with Nav1.7 fully in the resting state (holding voltage set to −140 mV) and on the same cells with the holding voltage set to produce about 20% inactivation. Compounds were not applied for a prespecified time, but for as long as required to achieve a stable level of block.
Measurement: IC50 = 2090 nM (pChEMBL 5.679999828338623)
Descriptors: MW 376.46, ALogP 3.41, HBA 6, HBD 3, PSA 91.83, QED 0.56
Compound (SMILES): O=S(=O)(Nc1ncccn1)c1ccc(NC(=S)N2CCN(c3ncnc4c3oc3ccccc34)CC2)cc1
InChIKey: OJMFWYZTRLXJOX-UHFFFAOYSA-N
Target: Mast/stem cell growth factor receptor Kit (SINGLE PROTEIN), Homo sapiens
UniProt: P10721
Assay: Inhibition of c-kit gain of function mutant in human GIST882 cells measured after 4 days by sulforhodamine B assay
Measurement: IC50 = 27500 nM (pChEMBL 4.559999942779541)
Descriptors: MW 546.64, ALogP 3.49, HBA 9, HBD 2, PSA 129.38, QED 0.31
Compound (SMILES): CNCC(O)CN(c1cc(OC)cc(OC)c1)c1ccc2ncc(-c3cnn(C)c3)nc2c1.Cl
InChIKey: VGBVCGQFMOSJMS-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 448.53, ALogP 2.77, HBA 9, HBD 2, PSA 97.56, QED 0.4
Compound (SMILES): COc1cc2c(Nc3ccc(Cl)c(Cl)c3)ncnc2cc1OCC1CN(c2ccncc2)CCO1
InChIKey: GBWHKHICVCJEQW-UHFFFAOYSA-N
Target: Ephrin type-A receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29317
Assay: Inhibition of recombinant human N-terminal His6-tagged EphA2 (N598 to R890 residues) expressed in baculovirus using poly-EY as substrate after 180 mins by luciferase coupled chemiluminescence assay
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 512.4, ALogP 5.37, HBA 8, HBD 1, PSA 81.63, QED 0.35
Compound (SMILES): COCCOCCSc1ccc(S(=O)(=O)NC(=O)c2ccc(N3CCN(CC4=C(c5ccc(Cl)cc5)CC(C)(C)CC4)CC3)cc2Oc2cccc3[nH]ccc23)cc1[N+](=O)[O-]
InChIKey: CDIIGXXJKVUCPX-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The inhibition constant (Ki) is the dissociation constant of an enzyme-inhibitor complex or a protein/small molecule complex, wherein the small molecule is inhibiting binding of one protein to another protein or peptide. Where the Ki for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-XL) is greater than the limits of detection of the assay used. Where the binding selectivity ratio for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the selectivity of a particular compound for Bcl-2 over Bcl-XL is at least as great as the number indicated. Where the Ki for a compound is represented as < (less than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-2) is lower than the limit of detection of the assay used. Inhibition constants were determined using Wang's equation (Wang Z-X).
Measurement: Ki = 24.21 nM (pChEMBL 7.619999885559082)
Descriptors: MW 888.51, ALogP 9.18, HBA 11, HBD 2, PSA 156.34, QED 0.04
Compound (SMILES): O=C(O)c1ccc(NC(=O)C(Cc2cccnc2)n2ccc(-c3cc(Cl)ccc3C(F)(F)F)cc2=O)cc1
InChIKey: LODANOLKNFDNSD-UHFFFAOYSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
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 of Tris/HCl pH 7.4; 100 mM of sodium chloride solution; 5 mM of calcium chloride solution; 0.1% of bovine serum albumin) and 20 μl of factor XIa from Kordia (0.45 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 factor XIa substrate Boc-Glu(OBzl)-Ala-Arg-AMC dissolved in assay buffer (10 μM 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 = 49 nM (pChEMBL 7.309999942779541)
Descriptors: MW 541.91, ALogP 5.7, HBA 5, HBD 2, PSA 101.29, QED 0.31
Compound (SMILES): N#CCC1(n2cc(-c3ccnc4[nH]ccc34)cn2)CN(C2CCN(C(=O)Nc3ccc(F)cc3C(F)(F)F)CC2)C1
InChIKey: CEQNCOJTCZYRDY-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Inhibition of human N-terminal His-tagged JAK2 catalytic domain (828 to 1132 residues) expressed in baculovirus infected insect cells assessed as decrease in phosphorylation of biotinylated peptide substrate after 1 hr in presence of 1 mM ATP by HTRF method
Measurement: IC50 > 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 566.56, ALogP 5.21, HBA 6, HBD 2, PSA 105.87, QED 0.32
Compound (SMILES): CNC(=O)c1c(-c2ccc(F)cc2)oc2cc(N(C)S(C)(=O)=O)c(-c3cnc4c(c3)-c3cc5ccccc5n3CO4)cc12
InChIKey: ACSHJRFZBHEKPQ-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1b (isolate Japanese) (HCV)
UniProt: P26662
Assay: ELISA-Based Assay: Measurement of inhibition by compounds was performed using the HCV replicon system. Several different replicons encoding different HCV genotypes or mutations were used. In addition, potency measurements were made using different formats of the replicon assay, including different ways of measurements and different plating formats. See Jan M. Vrolijk et al., A replicons-based bioassay for the measurement of interferons in patients with chronic hepatitis C, 110 J. VIROLOGICAL METHODS 201 (2003); Steven S. Carroll et al., Inhibition of Hepatitis C Virus RNA Replication by 2'-Modified Nucleoside Analogs, 278(14) J. BIOLOGICAL CHEMISTRY 11979 (2003). However, the underlying principles are common to all of these determinations, and are outlined below.Stable neomycin phosphotransferase encoding replicons-harboring cell lines were used, so all cell lines were maintained under G418 selection prior to the assay. Potency was determined using a cell ELISA assay with an antibody to the replicons encoded NS3/4a protease. See Caterina Trozzi et al., In Vitro Selection and Characterization of Hepatitis C Virus Serine Protease Variants Resistant to an Active-Site Peptide Inhibitor, 77(6) J. Virol. 3669 (2003).
Measurement: IC50 = 5.1 nM (pChEMBL 8.289999961853027)
Descriptors: MW 596.64, ALogP 6.03, HBA 7, HBD 1, PSA 106.67, QED 0.26
Compound (SMILES): N#CCC1(n2cc(-c3ncnc4[nH]ccc34)cn2)CN(C2CCN(C(=O)c3ccc(-c4cccc5ncccc45)c(F)c3)CC2)C1
InChIKey: WHGDXCBTRUREHN-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: In Vitro JAK Kinase Assay: Compounds herein were tested for inhibitory activity of JAK targets according to the following in vitro assay described in Park et al., Analytical Biochemistry 1999, 269, 94-104. The catalytic domains of human JAK1 (a.a. 837-1142), Jak2 (a.a. 828-1132) and Jak3 (a.a. 781-1124) with an N-terminal His tag were expressed using baculovirus in insect cells and purified. The catalytic activity of JAK1, JAK2 or JAK3 was assayed by measuring the phosphorylation of a biotinylated peptide. The phosphorylated peptide was detected by homogenous time resolved fluorescence (HTRF). IC50s of compounds were measured for each kinase in the 40 microL reactions that contain the enzyme, ATP and 500 nM peptide in 50 mM Tris (pH 7.8) buffer with 100 mM NaCl, 5 mM DTT, and 0.1 mg/mL (0.01%) BSA. For the 1 mM IC50 measurements, ATP concentration in the reactions was 1 mM. Reactions were carried out at room temperature for 1 hr and then stopped with 20 μL 45 mM EDTA, 300 nM SA-APC, 6 nM Eu-Py20 in assay buffer (Perkin Elmer, Boston, Mass.). Binding to the Europium labeled antibody took place for 40 minutes and HTRF signal was measured on a Fusion plate reader (Perkin Elmer, Boston, Mass.).
Measurement: IC50 < 5 nM (pChEMBL 8.300999641418457)
Descriptors: MW 611.69, ALogP 5.41, HBA 8, HBD 1, PSA 119.62, QED 0.27
Compound (SMILES): CCc1cc(C(F)(F)F)cnc1SCC(=O)N1CCCc2cccc(C)c21
InChIKey: IKSGEVCSNCNGJM-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8NET8
Assay: Inhibition Assay: To determine whether compounds were selective for TRPV3 inhibition over inhibition of other ion channel types, the human ERG (hERG), NaV1.2, and TRPV1 (hTRPV1) channels and the rat TRPV6 (rTRPV6) channel were also stably transfected and expressed or induced to express in mammalian cell lines. The methods for measuring currents from these channels are well-established and have been described in numerous publications (See, Weerapura et al., 2002, J Physiology 540: 15-27; Rush et al., 2005, J Physiology 564: 808-815; Caterina et al., 1997, Nature 389: 816-824; Hoenderhop et al., 2001, J Physiology 537: 747-761; Clapham et al., 2003, Pharmacol Rev 55: 591-596).
Measurement: IC50 < 5000 nM (pChEMBL 5.301000118255615)
Descriptors: MW 394.46, ALogP 5.04, HBA 3, HBD 0, PSA 33.2, QED 0.68
Compound (SMILES): CC1=C(N2CCC3(CCN(CC(O)c4ccc5c(c4Cl)COC5=O)CC3)C2=O)COC1=O
InChIKey: NPPNFYKHYXQRCK-UHFFFAOYSA-N
Target: ATP-sensitive inward rectifier potassium channel 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P48048
Assay: Thallium Flux Assay: The ROMK channel functional thallium flux assay is performed in 384 wells, using the FLIPR-Tetra instrument. HEK-hKir1.1 cells are seeded in Poly-D-Lysine microplates and kept in a 37° C.-10% CO2 incubator overnight. On the day of the experiment, the growth media is replaced with the FluxOR reagent loading buffer and incubated, protected from light, at ambient temperature (23-25° C.) for 90 min. The loading buffer is replaced with assay buffer+/-test compound followed by 30 min incubation at ambient temperature, where the Thallium/Potassium stimulant is added to the microplate.Step Protocol 1. Seed HEK-hKir1.1 cells (50 uA at 20,000 cells/well) in 384-well PDL coated Microplates 2. Allow cells to adhere overnight in humidified 37° C./10% CO2 incubator 3. Completely remove cell growth media from microplate and replace with 25 ul loading buffer 4. Incubate Microplate at room temperature, protected form light, for 90 min.
Measurement: IC50 = 10 nM (pChEMBL 8)
Descriptors: MW 460.91, ALogP 2.19, HBA 7, HBD 1, PSA 96.38, QED 0.69
Compound (SMILES): COCCOCC(C)(C)NC(=O)c1ccc(N2CC(F)(F)C2)c(-c2cccc(Cl)c2)n1
InChIKey: KFLVXDWUNBIPBE-UHFFFAOYSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1.times.HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37° C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30.degree. C. for 30 min Compounds were added to a final assay volume of 100 ul and incubated for 30 min at 30° C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 ul lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN.sub.3) and 50 ul detection solutions (20 uM mAb Alexa700-cAMP 1:1, and 48 uM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 453.92, ALogP 4.03, HBA 5, HBD 1, PSA 63.69, QED 0.58
Compound (SMILES): COc1cccc2c1NC(=O)[C@@H](NC(=O)[C@H](CCC(F)(F)F)[C@H](CCC(F)(F)F)C(N)=O)N=C2c1cccc(Cl)c1
InChIKey: BQJHUPVEKBVMDC-WJONJSRFSA-N
Target: Neurogenic locus notch homolog protein 3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UM47
Assay: Biological Assay: The pharmacological properties of the compounds of this invention may be confirmed by a number of biological assays.
Measurement: IC50 = 3.4 nM (pChEMBL 8.470000267028809)
Descriptors: MW 606.95, ALogP 4.98, HBA 5, HBD 3, PSA 122.88, QED 0.33
Compound (SMILES): COc1cc(-c2cc(F)cc(F)c2)c(C)cc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: ORNDBCORCUTKLC-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 = 1988 nM (pChEMBL 5.699999809265137)
Descriptors: MW 523.52, ALogP 5.04, HBA 7, HBD 1, PSA 103.43, QED 0.33
Compound (SMILES): O=C(O)CCc1ccc2cccc(N3CCN(CCc4ccc(OCCCN5CCCCCC5)cc4)CC3)c2n1
InChIKey: RRMGPLJQTAJCCP-UHFFFAOYSA-N
Target: Histamine H3 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5N1
Assay: Antagonist activity at human Histamine H3 receptor expressed in CHO cell membranes assessed as inhibition of histamine-induced [35S]GTPgammaS binding after 2 to 6 hrs by scintillation proximity assay
Measurement: Ki = 1.585 nM (pChEMBL 8.800000190734863)
Descriptors: MW 544.74, ALogP 5.26, HBA 6, HBD 1, PSA 69.14, QED 0.31
Compound (SMILES): O=S(=O)(Nc1nccs1)c1ccc2c(c1)OCCN2c1ccc(OC(F)(F)C(F)F)cc1
InChIKey: OPZAQYSOXZKYOS-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: 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 = 4720 nM (pChEMBL 5.329999923706055)
Descriptors: MW 489.47, ALogP 4.71, HBA 7, HBD 1, PSA 80.76, QED 0.48
Compound (SMILES): Cc1cc2ccc1[C@@H](C)COC(=O)Nc1ccc(OC(F)(F)F)c(c1)CN(C)C(=O)[C@@H]2Nc1ccc2c(N)nccc2c1
InChIKey: YLICDVIPJRKVDF-XRHLQHRESA-N
Target: Prothrombin (SINGLE PROTEIN), Homo sapiens
UniProt: P00734
Assay: Inhibition of alpha-thrombin (unknown origin) using pyroGlu-Pro-Arg-pNA as substrate assessed as release of pNA after 10 to 120 mins by spectrophotometric method
Measurement: Ki > 10000 nM (pChEMBL 5)
Descriptors: MW 593.61, ALogP 6.5, HBA 7, HBD 3, PSA 118.81, QED 0.25
Compound (SMILES): Cn1c(=O)n(C2CCOCC2)c2c3cc(-c4cnc(N)nc4)ccc3ncc21
InChIKey: YMLAOBUVHWUIQH-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase ATR (SINGLE PROTEIN), Homo sapiens
UniProt: Q13535
Assay: Cellular Assay: Assay b) HT29 cells (ECACC #85061109) were seeded into 384 well assay plates (Costar #3712) at a density of 6000 cells/well in 40 μl EMEM medium containing 1% L glutamine and 10% FBS and allowed to adhere overnight. The following morning compound of Formula (I) in 100% DMSO was added to assay plates by acoustic dispensing. After 1 h incubation at 37° C. and 5% CO2, 40 nl of 3 mM 4NQO in 100% DMSO was added to all wells by acoustic dispensing, except minimum control wells which were left untreated with 4NQO to generate a null response control. Plates were returned to the incubator for a further 1 h. Then cells were fixed by adding 20 μl of 3.7% formaldehyde in PBS solution and incubating for 20 mins at r.t. Then 20 μl of 0.1% Triton X100 in PBS was added and incubated for 10 minutes at r.t., to permeabalise cells. Then the plates were washed once with 50 μl/well PBS, using a Biotek EL405 plate washer. Phospho-Chk1 Ser 345 antibody (Cell Signalling Technology #2348) was diluted 150 fold in PBS containing 0.05% polysorbate/Tween and 15 μl was added to each well and incubated over night at r.t. The next morning plates were washed three times with 50 μl/well PBS, using a Biotek EL405 plate washer, and then 20 μl of secondary Ab solution, containing 500 fold diluted Alexa Fluor 488 Goat anti-rabbit IgG (Molecular Probes #A-11008) and 0.002 mg/ml Hoeschst dye (Molecular Probes #H-3570), in PBST, was added. After 2 h incubation at r.t., the plates were washed three times with 50 μl/well PBS, using a Biotek EL405 plate washer, and plates were then sealed with black plate seals until read. Plates were read using an ArrayScan VTI instrument, using an XF53 filter with 10× objective. A two laser set up was used to analyse nuclear staining with Hoeschst (405 nm) and secondary antibody staining of pChk1 (488 nm).
Measurement: IC50 = 12.9 nM (pChEMBL 7.889999866485596)
Descriptors: MW 376.42, ALogP 2.28, HBA 8, HBD 1, PSA 100.85, QED 0.58
Compound (SMILES): Cc1nc2cccc(-c3cc4c([nH]3)[C@H](CO)NC4=O)c2nc1NC1(C)CC1
InChIKey: BMRHAWSJNXXRJI-HNNXBMFYSA-N
Target: Serine/threonine-protein kinase pim-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P1W9
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 = 1.82 nM (pChEMBL 8.739999771118164)
Descriptors: MW 363.42, ALogP 2.67, HBA 5, HBD 4, PSA 102.93, QED 0.57
Compound (SMILES): CN1Cc2c(Cl)cc(Cl)cc2[C@H](c2cccc(S(=O)(=O)NCCOCCOCCNC(=O)NCCC[N+](C)(CCCNC(=O)NCCOCCOCCNS(=O)(=O)c3cccc([C@@H]4CN(C)Cc5c(Cl)cc(Cl)cc54)c3)CCCNC(=O)NCCOCCOCCNS(=O)(=O)c3cccc([C@@H]4CN(C)Cc5c(Cl)cc(Cl)cc54)c3)c2)C1
InChIKey: WXCCSPRGGWAWEV-BQGKCZGUSA-O
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 = 7.943 nM (pChEMBL 8.100000381469727)
Descriptors: MW 1788.73
Compound (SMILES): COCCNCc1ccc(Nc2nccc(-c3cnc(N4CC[C@H](F)C4)c(C#N)c3)n2)cn1
InChIKey: PJFYONWFRCIFBJ-SFHVURJKSA-N
Target: Serine/threonine-protein kinase TBK1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UHD2
Assay: Time-Resolved Fluorescence-based Lanthascreen Assay: Kinase inhibition assays (10 μL) were performed at 20° C. in 384-well plate format. Compound IC50 values were determined at the apparent Km for ATP (20 μM) based on a radiometric assay (Invitrogen) using 8 or 10 point curves in duplicate. The final reaction conditions contained 400 nM fluorescein-IkBα substrate (DRHDSGLDSMKDE), 20 μM ATP, 2 nM or 8 nM IKKε or TBK1 kinase respectively, and 3% DMSO in kinase assay buffer consisting of 50 mM HEPES (pH 7.5), 10 mM MgCl, 1 mM EGTA, 0.01% Brij-35.Compound dilutions were prepared from 10 mM DMSO stocks by dilution into DMSO. Compound dilution series were further diluted in kinase assay buffer to give a 12% DMSO stock, the final concentration in the assay being 3% DMSO.The kinase phosphorylation assay was initiated by the addition of the kinase and the reaction was allowed to proceed for 1 h or 2.5 h for IKKε and TBK-1 kinase respectively.
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 448.51, ALogP 2.83, HBA 9, HBD 2, PSA 111.88, QED 0.48
Compound (SMILES): NCCCCN(Cc1nccc2c3ccccc3n(CCCN)c12)[C@H]1CCCc2cccnc21
InChIKey: ANWJCECNNDZVCW-SANMLTNESA-N
Target: C-X-C chemokine receptor type 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P61073
Assay: Calcium Mobilization Assay: Functional modulation of CXCR4 was determined by calcium mobilization assay using leukemic lymphoid CEM cells, which naturally express high levels of CXCR4. Generally, the assay is carried out as follows: Cells are grown in vitro to confluency. On the day of the assay, cells are removed from the incubator, culture medium removed and replaced with a calcium sensitive dye (Calcium3 assay kit; Molecular Devices, Sunnyvale, Calif.). The cells are allowed to dye load for 45-60 min at 37 °C. in the cell culture incubator and then allowed to equilibrate to room temperature for no more than 15 min before assay. Compound plates were generated containing up to 3% dimethyl sulfoxide in media kept at room temperature. Test compounds were added to the cells at a 1:3 dilution, and calcium mobilization was measured using a Flex Station fluorescence imager (Molecular Devices). This first read was used to determine direct agonist activation of the CXCR4 receptor by the test compounds.
Measurement: IC50 = 7 nM (pChEMBL 8.149999618530273)
Descriptors: MW 456.64, ALogP 4.55, HBA 6, HBD 2, PSA 85.99, QED 0.34
Compound (SMILES): CCOC(=O)C(=O)Nc1ccc2nc(NC(=O)c3cc(Br)c(Br)[nH]3)sc2c1
InChIKey: QGKSDBJSDISIPL-UHFFFAOYSA-N
Target: DNA gyrase subunit B (SINGLE PROTEIN), Escherichia coli
UniProt: P0AES6
Assay: Inhibition of Escherichia coli DNA gyrase B ATP binding site assessed as reduction in supercoiling of relaxed pNO1 DNA after 30 mins in presence of biotinylated oligonucleotide by sybrGOLD staining based fluorescence assay
Measurement: IC50 = 29000 nM (pChEMBL 4.539999961853027)
Descriptors: MW 516.17, ALogP 3.9, HBA 6, HBD 3, PSA 113.18, QED 0.36