lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CNC(=O)c1c(-c2ccc(F)cc2)oc2cc(N(C)S(C)(=O)=O)c(-c3ccc4c(c3)-c3nc5ccccn5c3CO4)cc12
InChIKey: FOELMWBMOUGCTM-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 = 4 nM (pChEMBL 8.399999618530273)
Descriptors: MW 596.64, ALogP 5.87, HBA 7, HBD 1, PSA 106.15, QED 0.27
Compound (SMILES): Cc1ccc(C2=C(c3nn[nH]n3)C(=O)NC(c3ccc(OCCCCC(F)(F)F)cc3)(C(F)(F)F)C2)cc1
InChIKey: SQAAUQSVBJCIAT-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: MGAT LCMS Assay: The MGAT enzyme reactions were performed in CORNING Falcon 96-well Polypropylene plates, in a total volume of 60 uL of 50 mM Potassium Phosphate buffer pH 7.4, containing a final concentration of 100 uM 2-oleoylglycerol, 15 uM oleoyl-Coenzyme A and 0.0013 ug/uL Human or Mouse MGAT-2 or 0.0026 ug/uL Rat recombinant MGAT-2 membranes expressed in Sf9 cells. Assay plates were run through a fully automated robotics system and shaken for 5 seconds every minute for a total 10 minutes. The reactions were then quenched with 120 uL of ice cold methanol containing 1 ug/mL 1,2-distearoyl-rac-glycerol as the internal standard. Plates were shaken for 2 minutes and spun down to remove protein precipitation. After the spin, samples were transferred to LC/MS compatible PCR plates. For LC/MS analysis, a ThermoFisher Surveyor pump, utilizing a Waters Symmetry C8, 50x2.1 mm column, was used for the chromatography of enzyme products.
Measurement: IC50 = 112 nM (pChEMBL 6.949999809265137)
Descriptors: MW 539.48, ALogP 5.51, HBA 5, HBD 2, PSA 92.79, QED 0.29
Compound (SMILES): CN(C(=O)c1ccc(N2CC(F)(F)C2)c(OCC2CC2)n1)C1(C(F)(F)F)CC1
InChIKey: HGZFKHKPMYPBLX-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 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated as follows: FRET=T730-Alexa730-P(T645-B645) with P=Ru730-B730/Ru645-B645, where T730 is the test well measured at 730 nM, T645 is the test well measured at 645 nm, B730 and B645 are the buffer controls at 730 nm and 645 nm, respectively. cAMP content is determined from the function of a standard curve spanning from 10 μM to 0.13 nM cAMP.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 405.37, ALogP 3.49, HBA 4, HBD 0, PSA 45.67, QED 0.68
Compound (SMILES): Nc1ncc(-c2ccc(-n3cccn3)cc2)cc1C(=O)Nc1ccncc1
InChIKey: RRQPRTMYJUAVMC-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase TBK1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UHD2
Assay: TBK1 Cell Assay: Assay Overview: MDA-MB-468 cells are detached with HyQ-Tase, counted, and seeded into a 384-well clear bottom TC-surface plate at density of 10,000 cells per well in a total volume of 35 ul complete medium. Alternatively cells are directly seeded from frozen vials. Cells are pre-treated with inhibitor compounds for 1 h prior to Poly(I:C) stimulation. After 2 h of incubation with Poly(I:C), cells are fixed in (para)formaldehyde (PFA) and permeabilized with methanol (MeOH). The cells are then blocked and incubated with an anti-pIRF3 antibody at 4oC overnight. The primary antibody is washed off, an AlexaFluor488-conjugated secondary is added, cells are counterstained with propidium iodide followed by image acquisition on IMX Ultra high content reader.
Measurement: IC50 = 10000 nM (pChEMBL 5)
Descriptors: MW 356.39, ALogP 3.16, HBA 6, HBD 2, PSA 98.72, QED 0.59
Compound (SMILES): N#CC1(C(=O)N2C[C@H](CF)[C@H](Nc3c(C(N)=O)cnn4cc(-c5cnn(C6CC6)c5)nc34)C2)CC1
InChIKey: XOWRRBYTFORVLK-SUMWQHHRSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: Kinase Assay: The assay reactions were performed in U-bottom 384-well plates. The final assay volume was 30 μl prepared from 15 μl additions of enzyme and substrates (fluoresceinated peptide and ATP) and test compounds in assay buffer (20 mM HEPES pH 7.4, 10 mM MgCl2, 25 mM beta-glycerol phosphate, 0.015% Brij35 and 4 mM DTT). The reaction was initiated by the combination of GST-JAK1 enzyme with substrates and test compounds. The reactions were incubated at room temperature for 3 hours and terminated by adding 60 μl of 35 mM EDTA to each sample. Each reaction mixture was analyzed on the Caliper LABCHIP® 3000 (Caliper, Hopkinton, Mass.) by electrophoretic separation of the fluorescent substrate and phosphorylated product Inhibition data were calculated by comparison with no enzyme control reactions for 100% inhibition, and vehicle-only treated reactions for 0% inhibition. The final concentration of reagents in the assays was: ATP, 100 μM; fluoresceinated peptide, 1.5 μM; GST-JAK1, 12.5 nM; and DMSO, 1.6%.
Measurement: IC50 = 3.7 nM (pChEMBL 8.430000305175781)
Descriptors: MW 477.5, ALogP 1.54, HBA 9, HBD 2, PSA 147.23, QED 0.52
Compound (SMILES): FC(F)(F)Oc1cccc(Nc2ncc(Br)c(Nc3cc(C4CC4)n[nH]3)n2)c1
InChIKey: HCZLLRKMXVSZCP-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 >= 5000 nM (pChEMBL 5.301000118255615)
Descriptors: MW 455.24, ALogP 5.23, HBA 6, HBD 3, PSA 87.75, QED 0.47
Compound (SMILES): COC(=O)CCCC[S@](C)(=O)=NC(=O)c1cnc(N)c(C#Cc2cccc(NC(=O)c3occc3C)c2)c1
InChIKey: SXGZAYZKUGJQPT-LHEWISCISA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: VEGFR2 Cellular Assay: Automated FLIPR (Fluorometric Imaging Plate Reader) technology was used to screen for inhibitors of VEGF induced increases in intracellular calcium levels in fluorescent dye loaded endothelial cells. HUVEC (human umbilical vein endothelial cells) (Clonetics) were seeded in 384-well fibronectin coated black-walled plates overnight @ 37° C./5% CO2. Cells were loaded with calcium indicator Fluo-4 for 45 minutes at 37° C. Cells were washed 2 times (Elx405, Biotek Instruments) to remove extracellular dye. For screening, cells were pre-incubated with test agents for 30 minutes, at a single concentration (10 uM) or at concentrations ranging from 0.0001 to 10.0 uM followed by VEGF165 stimulation (10 ng/mL). Changes in fluorescence at 516 nm were measured simultaneously in all 384 wells using a cooled CCD camera. Data were generated by determining max-min fluorescence levels for unstimulated, stimulated, and drug treated samples.
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 536.61, ALogP 3.8, HBA 8, HBD 2, PSA 153.95, QED 0.25
Compound (SMILES): O=C(Cn1cnc(-c2ccccc2)c1-c1ccccc1)N1CCCCC1
InChIKey: XVWIFZWDNOIVMY-UHFFFAOYSA-N
Target: Prostaglandin G/H synthase 1 (SINGLE PROTEIN), Ovis aries
UniProt: P05979
Assay: Inhibition of ovine COX-1 assessed as reduction in oxidation of TMPD using arachidonic acid as substrate preincubated for 5 mins followed by substrate and TMPD addition measured after 5 mins by colorimetric assay
Measurement: IC50 = 8510 nM (pChEMBL 5.070000171661377)
Descriptors: MW 345.45, ALogP 4.23, HBA 3, HBD 0, PSA 38.13, QED 0.71
Compound (SMILES): Cc1cc(=O)oc2cc(NC(=O)[C@@H]3CCCN3C(=O)[C@H](CC(N)=O)NC(=O)[C@@H]3CCC(=O)N3)ccc12
InChIKey: YSNGYDACLOQMCZ-ULQDDVLXSA-N
Target: Thyrotropin-releasing hormone receptor (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q01717
Assay: Displacement of [3H]-3MeHis-TRH from TRH receptor in Sprague-Dawley rat cortical membranes
Measurement: Ki = 47863.01 nM (pChEMBL 4.320000171661377)
Descriptors: MW 497.51, ALogP -0.33, HBA 7, HBD 4, PSA 180.91, QED 0.38
Compound (SMILES): O=c1ccc2c(OCCCOc3ccccc3)c3ccoc3cc2o1
InChIKey: JTXGRXTXJLZYMG-UHFFFAOYSA-N
Target: Potassium voltage-gated channel subfamily A member 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22001
Assay: Inhibition of Kv1.3 (unknown origin)
Measurement: EC50 = 15 nM (pChEMBL 7.820000171661377)
Descriptors: MW 336.34, ALogP 4.39, HBA 5, HBD 0, PSA 61.81, QED 0.39
Compound (SMILES): O=C(O)c1ccc(COC[C@H]2CCC(=O)N2c2ccc(C(O)Cc3ccccc3)cc2)o1
InChIKey: UAXKGIZARLQFTR-PSDZMVHGSA-N
Target: Prostaglandin E2 receptor EP2 subtype (SINGLE PROTEIN), Homo sapiens
UniProt: P43116
Assay: Displacement of [3H]PGE2 from from human EP2 receptor expressed in HEK293 cells at pH 7.3 membranes incubated for 60 mins
Measurement: Ki = 2293 nM (pChEMBL 5.639999866485596)
Descriptors: MW 435.48, ALogP 3.97, HBA 5, HBD 2, PSA 100.21, QED 0.53
Compound (SMILES): C[C@H](Nc1nc(N)nc(N)c1C#N)c1nc2c(F)cc(F)cc2c(=O)n1-c1cnc(N)cn1
InChIKey: GFFQKKWMLSGONS-ZETCQYMHSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The activities were measured using a time-resolved fluorescence resonance energy transfer (TR-FRET) assay. TR-FRET monitored the formation of 3,4,5-inositol triphosphate molecule that competed with fluorescently labeled PIP3 for binding to the GRP-1 pleckstrin homology domain protein. An increase in phosphatidylinositide 3-phosphate product resulted in a decrease in TR-FRET signal as the labeled fluorophore was displaced from the GRP-1 protein binding site. 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 2xKm ATP (75-500 uM), 2 uM PIP2, 5% glycerol, 5 mM MgCl2, 50 mM NaCl, 0.05% (v/v) Chaps, 1 mM dithiothreitol, and 1% (v/v) DMSO at the following concentrations for each isoform: PI3Kα, PI3Kβ, and PI3Kδ between 25 and 50 uM, and PI3Kγ at 2 nM. The compounds of Table 1 and Compound X ((S)-2,4-diamino-6-((1-(5-chloro-4-oxo-3-(pyridin-3-yl)-3,4-dihydroquinazolin-2-yl)ethyl)amino)pyrimidine-5-carbonitrile) and Compound Y ((S)-2,4-diamino-6-((1-(5-chloro-4-oxo-3-(pyridin-3-yl)-3,4-dihydroquinazolin-2-yl)ethyl)amino)pyrimidine-5-carbonitrile) were added to the assay solution and incubated for 30 minutes at 25° C. The reactions were terminated with a final concentration of 10 mM EDTA, 10 nM labeled-PIP3, and 35 nM Europium labeled GRP-1 detector protein before reading TR-FRET on an Envision plate reader (Ex: 340 nm; Em: 615/665 nm; 100 us delay and 500 us read window). The results were normalized based on positive (1 uM wortmanin) and negative (DMSO) controls, and the IC50 values for PI3K α, β, δ and γ were calculated from the fit of the dose-response curves to a four-parameter equation.
Measurement: IC50 = 53 nM (pChEMBL 7.28000020980835)
Descriptors: MW 451.4, ALogP 1.04, HBA 12, HBD 4, PSA 200.33, QED 0.35
Compound (SMILES): O=C(CCCCCN1CCN(c2ccccc2-c2ccccc2)CC1)NCc1ccc(OCCCN2CCOCC2)cc1
InChIKey: HCQHYAXNIILCGJ-UHFFFAOYSA-N
Target: 5-hydroxytryptamine receptor 1A (SINGLE PROTEIN), Homo sapiens
UniProt: P08908
Assay: Displacement of [3H]-8-OH-DPAT from human 5-HT1A receptor expressed in HEK293 cell membranes after 1 hr by microbeta counting analysis
Measurement: Ki = 149 nM (pChEMBL 6.829999923706055)
Descriptors: MW 584.8, ALogP 5.45, HBA 6, HBD 1, PSA 57.28, QED 0.24
Compound (SMILES): Cn1c(=O)n([C@H]2CCCOC2)c2c3cc(-c4ccc(OCCCN5CCCC5)nc4F)ccc3ncc21
InChIKey: MQEUUOSLBCMAPQ-FQEVSTJZSA-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 > 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 505.59, ALogP 4.31, HBA 8, HBD 0, PSA 74.41, QED 0.28
Compound (SMILES): CC(C)n1ccc(C2=C(C(=O)NS(=O)(=O)C3CC3)C(=O)N[C@@](c3ccc(OCCCC(F)(F)F)cc3F)(C(F)(F)F)C2)n1
InChIKey: HNGZLXUUSVHYTC-DEOSSOPVSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: LCMS Assay: The MGAT enzyme reactions were performed in Corning FALCON® 96-well Polypropylene plates, in a total volume of 60 μL of 50 mM Potassium Phosphate buffer pH 7.4, containing a final concentration of 100 μM 2-oleoylglycerol, 15 μM oleoyl-Coenzyme A and 0.0013 μg/μL Human or Mouse MGAT-2 or 0.0026 μg/μL Rat recombinant MGAT-2 membranes expressed in Sf9 cells. Assay plates were run through a fully automated robotics system and shaken for 5 seconds every minute for a total 10 minutes. The reactions were then quenched with 120 μL of ice cold methanol containing 1 μg/mL 1,2-distearoyl-rac-glycerol as the internal standard. Plates were shaken for 2 minutes and spun down to remove protein precipitation. After the spin, samples were transferred to LC/MS compatible PCR plates. For LC/MS analysis, a ThermoFisher Surveyor pump, utilizing a Waters Symmetry C8, 50×2.1 mm column, was used for the chromatography of enzyme products. The buffer system consists of 0.1% formic acid in water with a mobile phase consisting 0.1% formic acid in methanol. The shallow gradient is 90-100% mobile phase in 0.2 min with a total run time of 2.3 min. The first 0.5 minutes of each injection was diverted to waste to eliminate the presence of Phosphate buffer in the enzymatic reaction. The column was run at 0.6 mL/min and a temperature of 65° C.
Measurement: IC50 = 35 nM (pChEMBL 7.460000038146973)
Descriptors: MW 640.58, ALogP 4.66, HBA 7, HBD 2, PSA 119.39, QED 0.22
Compound (SMILES): Cc1cncc(-c2cc3c(cn2)cnn3-c2cccc(N3CCC[C@H](O)C3)n2)n1
InChIKey: QEULBJUCNBXCDE-INIZCTEOSA-N
Target: Serine/threonine-protein kinase pim-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P1W9
Assay: Kinase Binding Assay: PIM-1, -2, and -3 enzymes were generated as fusion proteins expressed in bacteria and purified by IMAC column chromatography (Sun, X., Chiu, J. F., and He, Q. Y. (2005) Expert Rev. Proteomics, 2:649-657). A fluorescent-labeled Pim-specific peptide substrate, was custom synthesized by American Peptide Company (Sunnyvale, Calif.). Reaction Buffer contained 10 mM HEPES, pH 7.2, 10 mM MgCl2, 0.01% Tween 20, 2 mM DTT. Termination Buffer contained 190 mM HEPES, pH 7.2, 0.015% Brij-35, 0.2% Coating Reagent 3 (Caliper Life Sciences, Hopkinton, Mass.), 20 mM EDTA. Separation Buffer contained 100 mM HEPES, pH 7.2, 0.015% Brij-35, 0.1% Coating Reagent 3, 1:200 Coating Reagent 8 (Caliper Life Sciences, Hopkinton, Mass.), 10 mM EDTA and 5% DMSO.PIM reactions were carried out in a final volume of 10 uL per well in a 384-well plate. A standard enzymatic reaction, initiated by the addition of 5 uL 2Ã¿Â¿ATP and test compound to 5 uL of 2x enzyme and FAM-peptide, contained 20 uM PIM1, 50 uM PIM2, or 55 uM PIM3, 1 uM FAM-peptide, and 10 uM ATP, in Reaction Buffer. After 90 minutes of incubation at room temperature, the phosphorylation reaction was stopped by the addition of 10 uL Termination Buffer. The product and substrate in each independent reaction were separated on a 12-sipper microfluidic chip (Caliper Life Sciences, Hopkinton, Mass.) run on a Caliper LC3000 (Caliper Life Sciences, Hopkinton, Mass.). The separation of product and substrate was optimized by choosing voltages and pressure using Caliper's Optimizer software (Hopkinton, Mass.). The separation conditions used a downstream voltage of -500V, an upstream voltage of -2150V, and a screening pressure of -1.2 psi. The product and substrate fluorophore were excited at 488 nm and detected at 530 nm. Substrate conversion was calculated from the electropherogram using HTS Well Analyzer software (Caliper Life Sciences, Hopkinton, Mass.). Ki values for the test compound were calculated.
Measurement: Ki = 4.17 nM (pChEMBL 8.380000114440918)
Descriptors: MW 387.45, ALogP 2.54, HBA 8, HBD 1, PSA 92.85, QED 0.58
Compound (SMILES): O=C(O)c1nc2ccccc2n(C2CC3CCC(C2)N3C2CC3CCCC(C3)C2)c1=O
InChIKey: WKPMMJFYKKQWLO-UHFFFAOYSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: mu-Opioid Receptor Binding Assay: Radioligand dose-displacement binding assays for mu-opioid receptors used 0.2 nM[3H]-diprenorphine (NEN, Boston, Mass.), with 5-20 mg membrane protein/well in a final volume of 500 uL 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 5004, 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 uL/well).
Measurement: Ki = 3825 nM (pChEMBL 5.420000076293945)
Descriptors: MW 421.54, ALogP 4.23, HBA 5, HBD 1, PSA 75.43, QED 0.8
Compound (SMILES): COc1cc(-c2noc(CCCN(C)[C@H]3CC[C@H](OC(=O)C(O)(c4cccs4)c4cccs4)CC3)n2)c(Cl)cc1CNC[C@H](O)c1ccc(O)c2[nH]c(=O)ccc12
InChIKey: WFULHVJAURNUNM-ZDQHWEPJSA-N
Target: Muscarinic acetylcholine receptor M3 (SINGLE PROTEIN), Homo sapiens
UniProt: P20309
Assay: Muscarinic Receptor Binding Assay: The study of binding to human muscarinic M1, M2, M3, M4 and M5 receptors was performed using commercial membranes (Perkin Elmer) prepared from CHO-K1 cells. Radioligand binding experiments were conducted in 96 polypropylene well plates in a total volume of 200 ul. All reagents were dissolved in assay binding buffer (PBS with calcium and magnesium, SIGMA), except compounds that were dissolved in DMSO 100%. Non-specific binding (NSB) was measured in the presence of 1 uM atropine. [3H]-NMS was used as the radioligand at a concentration of 1 nM for M2, M3 and M5 and 0.3 nM for M1 and M4. [3H]-NMS and antagonists were incubated with membranes that express human muscarinic receptors M1, M2, M3, M4 and M5 at concentrations of 8.1, 10, 4.9, 4.5 and 4.9 ug/well, respectively. After an incubation period of two hours with gentle shaking, 150 ul of the reaction mix were transferred to 96 GF/C filter plates (Millipore), previously treated with wash buffer.
Measurement: IC50 = 0.2 nM (pChEMBL 9.699999809265137)
Descriptors: MW 834.42, ALogP 6.55, HBA 14, HBD 5, PSA 183.27, QED 0.07
Compound (SMILES): O=C1c2ccccc2C(O)(c2ccc3nc(Nc4ccccn4)[nH]c3c2)N1c1cccc(Cl)c1F
InChIKey: MUAXWDHYFWDBJL-UHFFFAOYSA-N
Target: RAF proto-oncogene serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P04049
Assay: Inhibition of c-Raf1 (unknown origin) pre-incubated for 30 mins by luciferase-luciferin-coupled chemiluminescence assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 485.91, ALogP 5.35, HBA 5, HBD 3, PSA 94.14, QED 0.32
Compound (SMILES): Cc1cc(F)ccc1Oc1cc(C(F)(F)F)ccc1C(=O)Nc1ccn(COP(=O)(O)O)c(=O)c1
InChIKey: SQDQNNKQTHOQHO-UHFFFAOYSA-N
Target: Sodium channel protein type 10 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Y9
Assay: Inhibition of human Nav1.8 expressed in HEK293 cells at -80 mV holding potential by whole-cell patch-clamp technique
Measurement: IC50 = 33.45 nM (pChEMBL 7.480000019073486)
Descriptors: MW 516.34, ALogP 4.43, HBA 6, HBD 3, PSA 127.09, QED 0.31
Compound (SMILES): O=C(c1ccc(N2CC(F)(F)C2)c(OCC2CC2)n1)N1CCCC12CCC2
InChIKey: FWLFSZKDWQACBC-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 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated as follows: FRET=T730-Alexa730-P(T645-B645) with P=Ru730-B730/Ru645-B645, where T730 is the test well measured at 730 nM, T645 is the test well measured at 645 nm, B730 and B645 are the buffer controls at 730 nm and 645 nm, respectively. cAMP content is determined from the function of a standard curve spanning from 10 μM to 0.13 nM cAMP.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 377.44, ALogP 3.48, HBA 4, HBD 0, PSA 45.67, QED 0.79
Compound (SMILES): FC(F)(F)COC1CN(C[C@H]2CC[C@H](c3nnn4cnc5[nH]ccc5c34)CC2)C1
InChIKey: WHVUOWTWWKFBGC-JOCQHMNTSA-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 = 170 nM (pChEMBL 6.769999980926514)
Descriptors: MW 408.43, ALogP 3.14, HBA 6, HBD 1, PSA 71.34, QED 0.7
Compound (SMILES): CN[C@@H](C)C(=O)N[C@@H]1C(=O)N(Cc2c(OC)ccc3cc(Br)ccc23)c2ccccc2O[C@@]1(C)COC
InChIKey: QZAXSBDPPGLMSO-VFQAHANWSA-N
Target: E3 ubiquitin-protein ligase XIAP (SINGLE PROTEIN), Homo sapiens
UniProt: P98170
Assay: TR-FRET Assay: Ten nanomolar of 6x Histidine-tagged BIR2 domain, corresponding to amino acids 124-240 of XIAP, or BIR3 domain, corresponding to amino acids 241-356 of XIAP, was mixed with 20 nM of the peptide AVPIAQKSEK-(8-biotin)-OH 1:2 TFA, in the presence of 50 mM Tris-C1, pH 7.5, 100 mM NaCl, 1 mM dithiothreitol (DTT) and 0.1 mg/mL bovine serum albumin (BSA). Following a 45 min. incubation at 37° C., Europium-Streptavidin and Allophycocyanin conjugated anti-Histidine antibody were added to a final concentration of 1.5 nM and 15 nM, respectively. Time-resolved fluorescence resonance energy transfer (TR-FRET) signals were measured 1 hour later at room temperature. Test compound potency was assessed at 10 serially diluted concentrations.
Measurement: IC50 = 25525 nM (pChEMBL 4.590000152587891)
Descriptors: MW 570.48, ALogP 4.03, HBA 6, HBD 2, PSA 89.13, QED 0.43
Compound (SMILES): COc1cc(N2CCC(O)CC2)ccc1Nc1nccc(-c2c(-c3cccc(C(=O)Nc4c(F)cccc4F)c3)nc3ccccn23)n1
InChIKey: KYVIQYWGGPLPCH-UHFFFAOYSA-N
Target: Insulin receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P06213
Assay: Inhibition of human GST-rTEV-tagged insulin receptor (979 to 1382 residues) expressed in baculovirus infected insect Sf9 cells assessed as decrease in biotin-aminohexyl-AEEEEYMMMMAKKKK-NH2 phosphorylation after 1 hr by fluorometer
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 647.69, ALogP 6.7, HBA 9, HBD 3, PSA 116.91, QED 0.17
Compound (SMILES): Nc1ncc(C2CCC(O)CC2)nc1-c1ccc(C(=O)NCc2ccccc2)c(F)c1
InChIKey: FFGOLDWBKVSWEI-UHFFFAOYSA-N
Target: Mitogen-activated protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P28482
Assay: Kinase Assay (New): Compound potency against activated ERK2 is determined using a kinase assay that measures ERK2-catalyzed phosphorylation of biotinylated ERKtide peptide substrate ([Biotin]-AHA-K-R-E-L-V-E-P-L-T-P-S-G-E-A-P-N-Q-A-L-L-R-[NH2], the peptide sequence derived from EGF receptor: SEQ ID NO:1). The assay is carried out in 50 mM HEPES [pH 7.5], 5 mM MgCl2, 1 mM DTT, 0.01% Tween-20, 0.05% BSA using 0.25 nM ERK2, 200 nM ERKtide peptide and 35 uM ATP (all concentrations are final in the reaction) in a total volume of 10.25 uL. A 16-point, half-log dilution series of compounds at 41x final concentration is used for generating IC50 curves. Compound dilution series are prepared in 100% DMSO. ERK2 is preincubated with compounds for 30 minutes at ambient temperature. Reaction is initiated by addition of a substrate cocktail of ERKtide peptide and ATP and is allowed to proceed for 2-3 hours at ambient temperature. Reaction is terminated by addition of 10 uL of a 2x stop buffer consisting of 100 mM Tris-Cl [pH 7.5], 25 mM EDTA, 0.01% Tween 20, 10 ug/mL of AlphaScreen Protein A Acceptor Beads, 10 ug/mL of Streptavidin Donor Beads (PerkinElmer, Waltham, Mass.), and 1.4 ug/mL phospho-EGF Receptor (Thr669) antibody (Cat #3056, Cell Signaling Technology, Danvers, Mass.). Terminated reactions are read, after overnight incubation in the dark, on an EnVision Multilabel Plate Reader (PerkinElmer, Waltham, Mass.), with excitation and emission wavelengths set to 680 nm and 570 nm, respectively. IC50 values are determined using a four-parameter fit.
Measurement: IC50 = 235 nM (pChEMBL 6.630000114440918)
Descriptors: MW 420.49, ALogP 3.81, HBA 5, HBD 3, PSA 101.13, QED 0.58
Compound (SMILES): O=C(N1CCCC[C@@H]1Cc1ccccc1)n1cc(-c2ccc(-c3ccccc3)cc2)nn1
InChIKey: JKJMWHULJIOKPJ-RUZDIDTESA-N
Target: Diacylglycerol lipase-alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y4D2
Assay: Inhibition of recombinant human DAGLalpha expressed in HEK293T cell membranes overexpressing MAGL using natural substrate 1-stearoyl-2-arachidonoly-sn-glycerol by fluorescence based assay
Measurement: IC50 = 25.12 nM (pChEMBL 7.599999904632568)
Descriptors: MW 422.53, ALogP 5.68, HBA 4, HBD 0, PSA 51.02, QED 0.43
Compound (SMILES): [C-]#[N+]c1c(Cl)cccc1CN1CCC(N2CCC(C(=O)c3ccc(OC)cc3)CC2)C1=O
InChIKey: XZVNTIRJCBONRF-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase tankyrase-1 (SINGLE PROTEIN), Homo sapiens
UniProt: O95271
Assay: Biochemical Assay: The autoparsylation activity of the TNKS 1/2 or PARP1/2 enzymes was measured by the liquid chromatography-mass spectrometry (LC/MS) detection of nicotinamide as readout. Compound activity in inhibiting the TNKS and PARP autoparsylation was evaluated by IC50 measurements. In the compound screening assays, the reaction is composed of 5 uL of compound in 8-point serial dilutions with concentrations ranging from 0.0086 to 18.75 uM, 20 nM of purified enzyme, and 250 uM of p-NAD+ in the 1x Assay Buffer. After 60 min incubation at room temperature, the reactions were quenched by the addition of 10 uL of 5x quenching solution (20% formic acid and 500 nM [d]-nicotinamide in water). For the background control wells, 10 uL of the 5x quenching solution per well was added prior to the addition of p-NAD+. The % Inhibition was calculated as: (Control-Sample)/(Control-Background)*100. "Control" is the average value of 8 wells without compound; and "Background" is the average of 8 wells mixed with 5x quenching solution measured prior to initiation of the reaction.
Measurement: IC50 = 162 nM (pChEMBL 6.789999961853027)
Descriptors: MW 451.95, ALogP 4.6, HBA 4, HBD 0, PSA 54.21, QED 0.48
Compound (SMILES): CC(C)c1nc2c(c(NCCc3ccccc3Cl)n1)CCN(c1ncccc1C(F)(F)F)CC2
InChIKey: ORSNFJDPYGXSED-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: O35433
Assay: Functional Assay: The assay was performed as described below, using HEK293 cells transfected with rat TRPV1 (rTRPV1/HEK293). These cells had a geneticin selection marker and were grown in Dulbecco's Modified Eagle Medium (DMEM, Gibco BRL) supplemented with 10% fetal bovine serum, penicillin/streptomycin (50 units/mL), and 500 μg/mL geneticin in 5% CO2 at 37° C. Cells were passaged frequently, every 3-5 days, to avoid overgrowth, depletion of essential medium components, or acidic medium exposure. Cells were passaged using a brief wash in 0.05% trypsin with 1 mM EDTA, followed by dissociation in divalent-free phosphate-buffered saline (Hyclone #SH30028.02). Dissociated cells were seeded onto poly-D-lysine coated black-walled 96-well plates (Biocoat; Becton Dickinson #354640) at about 40,000 cells per well and grown for approximately 1 day in culture medium to near confluency. The assay buffer was composed of 130 mM NaCl, 2 mM KCl, 2 mM MgCl2, 10 mM HEPES, 5 mM glucose, and either 2 mM or 20 μM CaCl2. On the day of the experiment, the culture medium was replaced with 2 mM calcium assay buffer using an automated plate washer (ELx405; Biotek, VT). The cells were incubated in 100 μL/well Fluo-3/AM (2 μM; TEFLabs #0116) with Pluronic F127 (100 μg/mL; Sigma #P2443) for 1 h at rt in the dark. After loading the cells, the dye solution was replaced with 50 μL/well of 20 μM calcium assay buffer using the ELx405 plate washer. Test compounds (50 μL/well) were added to the plate and incubated for 30 min. Intracellular Ca2+ levels were subsequently assayed using a Fluorometric Imaging Plate Reader (FLIPR™ instrument, Molecular Devices, CA) to simultaneously monitor Fluo-3 fluorescence in all wells (λexcitation=488 nm, λemission=540 nm) during challenge with agonist (capsaicin). The IC50 values were determined. Cells were challenged with 150 nM capsaicin and the fluorescence counts were captured following agonist addition at a sampling rate of 0.33 Hz. The contents of the wells were mixed 3 times (40 μL mix volume) immediately after the additions were made. Concentration dependence of block was determined by exposing each well of cells in duplicate rows of a 96 well plate to a serial dilution of test compound. The concentration series usually started at 10 Î¼M with a three-fold serial decrement in concentration. The magnitude of the capsaicin response was determined by measuring the change in fluo3 fluorescence before and 100 seconds after the addition of the agonist.
Measurement: IC50 = 1300 nM (pChEMBL 5.889999866485596)
Descriptors: MW 489.97, ALogP 5.93, HBA 5, HBD 1, PSA 53.94, QED 0.47
Compound (SMILES): Nc1ncnc(N2CCC(c3nc(-c4cccc(C(F)(F)F)c4)cn3CCN3CCCC3)CC2)c1Br
InChIKey: NLKBUXJOZSFKGH-UHFFFAOYSA-N
Target: Ribosomal protein S6 kinase beta-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23443
Assay: P70S6K Enzyme Assay: P70S6K inhibitor compounds are diluted and plated in 96 well plates. A reaction mixture including the following components is then added to the compound plate to initiate the enzyme reaction; P70S6K (3 nM, T412E mutant, Millipore) is mixed with 24 uM ATP in an assay buffer containing 100 mM Hepes (pH 7.5), 5 mM MgCl2, 1 mM DTT, 0.015% Brij and 1 uM of the substrate peptide FITC-AHA-AKRRRLSSLRA-OH (derived from the S6 ribosomal protein sequence, FITC=fluorescein isothiocyanate, AHA=6-aminohexanoic acid). The reaction is incubated for 90 min at 25.degree. C., before the addition of 10 mM EDTA to stop the reaction. The proportion of substrate and product (phosphorylated) peptide is analysed on a Caliper Life Sciences Lab Chip 3000, using a pressure of -1.4 psi, and upstream and downstream voltages of -3000 and -700 respectively. Product peaks are resolved before substrate peaks on the resulting chromatograms.
Measurement: IC50 = 2.2 nM (pChEMBL 8.65999984741211)
Descriptors: MW 564.45, ALogP 5.18, HBA 7, HBD 1, PSA 76.1, QED 0.44
Compound (SMILES): CCOC(=O)N=S(C)(=O)Cc1cccc(Nc2ncc(F)c(-c3ccc(F)cc3OC)n2)c1
InChIKey: VQAYFOJQHDLFNL-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P24941
Assay: CDK2/CycE Kinase Assay: Recombinant fusion proteins of GST and human CDK2 and of GST and human CycE, expressed in insect cells (Sf9) and purified by Glutathion-Sepharose affinity chromatography, were purchased from ProQinase GmbH (Freiburg, Germany). As substrate for the kinase reaction biotinylated peptide biotin-Ttds-YISPLKSPYKISEG (C-terminus in amid form) was used which can be purchased e.g. form the company JERINI Peptide Technologies (Berlin, Germany)For the assay 50 nl of a 100 fold concentrated 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 CDK2/CycE in aqueous assay buffer [50 mM Tris/HCl pH 8.0, 10 mM MgCl2, 1.0 mM dithiothreitol, 0.1 mM sodium ortho-vanadate, 0.01% (v/v) Nonidet-P40 (Sigma)] were added and the mixture was incubated for 15 min at 22 C. to allow pre-binding of the test compounds to the enzyme before the start of the kinase reaction.
Measurement: IC50 = 780 nM (pChEMBL 6.110000133514404)
Descriptors: MW 476.51, ALogP 4.93, HBA 7, HBD 1, PSA 102.77, QED 0.52
Compound (SMILES): Cc1c(Nc2ccccc2)nnc(N2CCN(c3cnc(C(C)(C)O)cn3)[C@H](C)C2)c1C
InChIKey: PTRDTDXUYJZTNM-MRXNPFEDSA-N
Target: Protein smoothened (SINGLE PROTEIN), Mus musculus
UniProt: P56726
Assay: Inhibition of mouse recombinant Shh protein in mouse TMHh12 cells assessed as inhibition of 25 nM Smo agonist-induced hedgehog signalling response by Gli-luciferase reporter gene assay
Measurement: IC50 = 100 nM (pChEMBL 7)
Descriptors: MW 433.56, ALogP 3.57, HBA 8, HBD 2, PSA 90.3, QED 0.63
Compound (SMILES): Cc1c[nH]c2ncc(-c3cccc(NC(=O)/C=C/CN(C)C)c3)cc12
InChIKey: POFBZQXDWPOYEW-VMPITWQZSA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
Assay: In Vitro Inhibition Assay: ITK and JAK3 Kinase assay procedures:Enzyme was incubated with substrate peptide in reaction buffer in the presence and absence of test compounds or Staurosporine. All additions were done on ice, followed by the addition of ATP mix. Wells were uniformly mixed using an Eppendorff plate shaker and incubated at 30° C. for 20 min, and stopped by the addition of 5 μL of 3% phosphoric acid. Volume was increased to 100 μL by adding 0.8% phosphoric acid which was then transferred to PC filter mats (Millipore), pre-equilibrated with 70% ethanol and water. Plates were washed thrice with 100 μL 0.8% phosphoric acid and dried for an hour at 60° C. 100 μL scintillation fluid was added into each well and reading taken in Perkin Elmer TOPCOUNT beta counter. The data analysis was performed by averaging the duplicate top count readings for each standard, negative, positive control (enzyme control) and samples and subtracting the average negative control from each reading which results in corrected values.
Measurement: IC50 > 100 nM (pChEMBL 7)
Descriptors: MW 334.42, ALogP 3.59, HBA 3, HBD 2, PSA 61.02, QED 0.7
Compound (SMILES): O=C(Nc1ccc(Cl)c(S(=O)(=O)C2CCCNC2)c1O)Nc1cccc(F)c1Cl
InChIKey: ZVDORRYGKFXKJT-UHFFFAOYSA-N
Target: C-X-C chemokine receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P25025
Assay: Displacement of [125I]IL8 from recombinant human CXCR2 expressed in CHO cell membranes measured after 1 hr by TopCount scintillation counting method
Measurement: IC50 < 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 462.33, ALogP 4.01, HBA 5, HBD 4, PSA 107.53, QED 0.51
Compound (SMILES): N#Cc1cc(-c2ccnc(Nc3ccc(OCCN4CC[C@H](F)C4)nc3)n2)cnc1N1CC[C@H](F)C1
InChIKey: PRZNLTVMMUQQBI-PMACEKPBSA-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 492.53, ALogP 3.52, HBA 9, HBD 1, PSA 103.09, QED 0.51
Compound (SMILES): CC(C)[C@H](NC(=O)C(=O)Nc1cccc2ccccc12)C(=O)N[C@@H](CC(=O)O)C(=O)COc1ccc(O)cc1
InChIKey: HIAYBHWMULSKJU-DHLKQENFSA-N
Target: Caspase-3 (SINGLE PROTEIN), Homo sapiens
UniProt: P42574
Assay: Inhibition of recombinant CPP32 (unknown origin) pre-incubated for 60 mins with Acetyl-Tyr-Val-Ala-Asp-amino-4-methylcoumarin substrate followed by enzyme addition by fluorescence based assay
Measurement: Ki = 42 nM (pChEMBL 7.380000114440918)
Descriptors: MW 535.55, ALogP 2.23, HBA 7, HBD 5, PSA 171.13, QED 0.23
Compound (SMILES): CCCN(CC1CC1)c1ccc(S(=O)(=O)NCC2CCOCC2)c(C(F)(F)F)c1
InChIKey: KGLLFFYVUDKJPE-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: Binding Assay: The binding assays were performed by testing the ability of the new compounds to displace the radiolabeled synthetic non-selective cannabinoid agonist [3H]CP55940 (168 Ci/mmol; PerkinElmer) from the human CB1 (hCB1) or human CB2 (hCB2) receptors on membranes derived from stably transfected HEK-293 cells (PerkinElmer). Membranes were diluted in assay buffer (50 mM Tris-HCl, 1 mM EDTA, 1 mM MgCl2, 1 mg/ml BSA, pH=7.4). The amount of membrane was determined for each batch of membranes according to protein binding assay. The minimum amount of membrane that gave 50% specific binding was used for the binding assay. In most assays, binding was tested using 8 μg and 4 μg protein of hCB1 and hCB2 membranes, respectively. The tested compounds were dissolved in DMSO and diluted in assay buffer to a final concentration of 0.1% solvent. Total binding of [3H]CP55940 was evaluated with 1.5 nM to hCB1 and with 0.5 nM to hCB2, according to Kd affinity of [3H]CP55940 for the respective membranes. The ability of the tested compounds to displace [3H]CP55940 was evaluated first at a single concentration point of 100 nM for binding toward hCB2 or hCB1. In certain cases, the displacement was tested at compound concentrations ranging from 0.03 nM to 10 μM. Non-specific binding was measured by the addition of 1 μM of unlabelled CP55940 to the tubes. Binding assays were performed in triplicate in a total volume of 500 μl for 60 minutes at 30° C., in a shaking bath. Free and bound radioligands were separated by rapid filtration through GF/C filter plates (PerkinElmer) that had been presoaked with 0.1% Polyethylenimine (Sigma). Filters were shaken for 1 hour in 7 ml scintillation fluid (PerkinElmer) and radioactivity was determined by liquid scintillation counter (Wallac; PerkinElmer).
Measurement: IC50 = 743 nM (pChEMBL 6.130000114440918)
Descriptors: MW 434.52, ALogP 4.04, HBA 4, HBD 1, PSA 58.64, QED 0.64
Compound (SMILES): CCCN[C@H](Cc1ccc(OCC)cc1)C(=O)N(C)[C@@H](Cc1ccccc1)C(=O)NCc1ccc(CN)cc1
InChIKey: RRCQNIGKGMQKJC-IHLOFXLRSA-N
Target: Kallikrein-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P06870
Assay: In Vitro Inhibition Assay: KLK1 inhibitory activity in vitro was determined using standard published methods (see e.g. Johansen et al., Int. J. Tiss. Reac. 1986, 8, 185; Shori et al., Biochem. Pharmacol., 1992, 43, 1209; Stürzebecher et al., Biol. Chem. Hoppe-Seyler, 1992, 373, 1025). Human KLK1 (Callbiochem) was incubated at 37° C. with the fluorogenic substrate H-DVal-Leu-Arg-AFC and various concentrations of the test compound. Residual enzyme activity (initial rate of reaction) was determined by measuring the change in optical absorbance at 410 nm and the IC50 value for the test compound was determined.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 530.71, ALogP 3.84, HBA 5, HBD 3, PSA 96.69, QED 0.28
Compound (SMILES): COC(=O)C12CC3CC(C1)C(OC(=O)N1CCC(c4ncc(-c5ccn(C)c(=O)c5)cn4)C1)C(C3)C2
InChIKey: UHDSCLOMXJMJPL-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 > 100 nM (pChEMBL 7)
Descriptors: MW 492.58, ALogP 3.14, HBA 8, HBD 0, PSA 103.62, QED 0.6
Compound (SMILES): COc1ccc2ccccc2c1[C@@H]1C[C@H](F)C(=O)N1Cc1ccc(OC(F)(F)F)cc1
InChIKey: JVNMWFIXSFYSEB-OALUTQOASA-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 = 552 nM (pChEMBL 6.260000228881836)
Descriptors: MW 433.4, ALogP 5.56, HBA 3, HBD 0, PSA 38.77, QED 0.49