lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CNC(=N)NCCC[C@H](NC(=O)[C@H](CC(C)C)NC(=O)NNC(=O)[C@H](Cc1cccc(F)c1)NC(=O)[C@@H](NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CCC(=O)O)NC(=O)[C@@H](Cc1ccc(O)cc1)NC(C)=O)[C@@H](C)O)C(=O)N[C@@H](Cc1c[nH]c2ccccc12)C(N)=O
InChIKey: BNSPMRCFHAFQER-WBDBVSIISA-N
Target: KiSS-1 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q969F8
Assay: Agonist activity at human KISS1R expressed in CHO/dhfr cells assessed as increase in intracellular Ca2+ levels measured for 180 secs by Fluo 3-AM dye based FLIPR assay
Measurement: EC50 = 0.47 nM (pChEMBL 9.329999923706055)
Descriptors: MW 1259.36
Compound (SMILES): Cc1ccc(C2=C(C(=O)Nc3cc(C)[nH]n3)C(=O)NC(c3ccc(OCCC(F)(F)F)cc3)(C(F)(F)F)C2)cc1
InChIKey: HCNHJVGAIPYXLH-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 = 1050 nM (pChEMBL 5.980000019073486)
Descriptors: MW 566.5, ALogP 5.73, HBA 4, HBD 3, PSA 96.11, QED 0.25
Compound (SMILES): CN(C)C(=O)CCCN(C)C1CCN(C(=O)Cn2cc(NC(=O)c3cnn4cccnc34)c(-c3cc(Cl)ccc3OC(F)F)n2)CC1
InChIKey: XVNALBBKCPINNQ-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: JAK Enzyme Assay: The activity of the isolated recombinant JAK1 and JAK2 kinase domain was measured by monitoring phosphorylation of a peptide derived from JAK3 (Val-Ala-Leu-Val-Asp-Gly-Tyr-Phe-Arg-Leu-Thr-Thr, fluorescently labeled on the N-terminus with 5-carboxyfluorescein) using the Caliper LabChip® technology (Caliper Life Sciences, Hopkinton, Mass.). To determine inhibition constants (Ki), compounds were diluted serially in DMSO and added to 50 μL kinase reactions containing purified enzyme (1.5 nM JAK1, or 0.2 nM JAK2), 100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 1.5 μM peptide substrate, ATP (25 μM), 10 mM MgCl2, 4 mM DTT at a final DMSO concentration of 2%. Reactions were incubated at 22° C. in 384-well polypropylene microtiter plates for 30 minutes and then stopped by addition of 25 μL of an EDTA containing solution (100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 150 mM EDTA), resulting in a final EDTA concentration of 50 mM. After termination of the kinase reaction, the proportion of phosphorylated product was determined as a fraction of total peptide substrate using the Caliper LabChip® 3000 according to the manufacturer's specifications.
Measurement: Ki = 1.72 nM (pChEMBL 8.760000228881836)
Descriptors: MW 672.14, ALogP 3.89, HBA 10, HBD 1, PSA 130.2, QED 0.24
Compound (SMILES): CCN1CCN(c2noc(COc3cc4ncnc(Nc5ccc(Cl)c(Cl)c5)c4cc3OC)n2)CC1
InChIKey: FGHXOSBHGQKMBX-UHFFFAOYSA-N
Target: Ephrin type-B receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P54760
Assay: Inhibition of recombinant human N-terminal His6-tagged EphB4 kinase domain (E605 to E890 residues) expressed in baculovirus using poly-AEKY as substrate after 150 mins in presence of gamma33P-ATP by liquid scintillation spectrometry
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 530.42, ALogP 4.79, HBA 10, HBD 1, PSA 101.67, QED 0.34
Compound (SMILES): O=C(Nc1ccc(C2CCCCN2)cc1)c1ccc(Cl)cc1
InChIKey: ZEDVYTYZHLXBDQ-UHFFFAOYSA-N
Target: Trace amine-associated receptor 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q923Y8
Assay: Radioligand Binding Assay: HEK-293 cells stably expressing mouse 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 = 6378.2 nM (pChEMBL 5.199999809265137)
Descriptors: MW 314.82, ALogP 4.41, HBA 2, HBD 2, PSA 41.13, QED 0.88
Compound (SMILES): Cc1nnc(-c2cc(C(=O)N3CC(c4ccc(C#N)cc4)C3)ccc2C2CCC2)[nH]1
InChIKey: PIISJIUUAMMZEV-UHFFFAOYSA-N
Target: Fatty acid synthase (SINGLE PROTEIN), Homo sapiens
UniProt: P49327
Assay: Biochemical Assay: The FASN enzyme was isolated from SKBr3 cells. SKBr3 is a human breast cancer cell-line with high levels of FASN expression. It is estimated that FASN comprises about 25% of the cytosolic proteins in this cell line. SKBr3 cells were homogenized in a dounce homogenizer then centrifuged for 15 minutes at 4° C. to remove particulate matter. The supernatant was then analyzed for protein content, diluted to the appropriate concentration, and used to measure FASN activity. The presence of FASN was confirmed by western blot analysis. A similar method for isolation of FASN from SKBr3 cells is described in Teresa, P. et al. (Clin. Cancer Res. 2009; 15(24), 7608-7615). FASN activity of the SKBr3 cell extract was determined by measuring either NADPH oxidation or the amount of thiol-containing coenzyme A (CoA) released during the fatty acid synthase reaction. The dye CPM (7-diethylamino-3-(4'-maleimidyl-phenyl)-4-methylcoumarin) contains a thiol reactive group that increases its fluorescence emission on reaction with the sulfhydryl group of CoA.
Measurement: IC50 = 97 nM (pChEMBL 7.010000228881836)
Descriptors: MW 397.48, ALogP 4.16, HBA 4, HBD 1, PSA 85.67, QED 0.72
Compound (SMILES): C[C@H](NC(=O)C(F)F)c1ccc(CN2CCc3cc(OCC4CC4(F)F)ccc3C2)cc1
InChIKey: ZNSIZHQNFVOQTM-OOJLDXBWSA-N
Target: Acetyl-CoA carboxylase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O00763
Assay: Spectrophotometric 384 Well Assay: Assay reactions are then carried out in 384-well plates, with hACC2 in an appropriate dilution and at final assay concentrations (f.c.) of 100 mM Tris (pH 7.5), 10 mM trisodium citrate, 25 mM KHCO3, 10 mM MgCl2, 0.5 mg/ml BSA, 3.75 mM reduced L-glutathione, 15 U/ml lactate dehydrogenase, 0.5 mM phosphoenolpyruvate, 15 U/ml pyruvate kinase, compounds at different concentrations at final DMSO concentrations of 1%.The enzymatic reaction is then started by addition of a mixture of NADH, acetyl Coenzyme A (both 2000 f.c.) and ATP (500 uM f.c.). The decrease of the optical density (slope S) is then determined at 25° C. at a wavelength of 340 nm over 15 minutes in a spectrophotometric reader.Each assay microtiter plate contains wells with vehicle instead of compound as controls for the non-inhibited enzyme (100% CTL; HIGH) and wells without acetyl-CoA as controls for non-specific NADH degradation (0% CTL; LOW).
Measurement: IC50 = 25 nM (pChEMBL 7.599999904632568)
Descriptors: MW 450.48, ALogP 4.72, HBA 3, HBD 1, PSA 41.57, QED 0.59
Compound (SMILES): O=C(Nc1cc(-c2ccc(F)c(NCC3CCOCC3)n2)c(Cl)cn1)[C@@H]1CCCNC1
InChIKey: OETJRQWLHMSDMD-OAHLLOKOSA-N
Target: CDK9/cyclin T1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P50750
Assay: Inhibition of CDK9/cyclin T1 (unknown origin) using cdk7tide peptide as substrate
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 447.94, ALogP 3.71, HBA 6, HBD 3, PSA 88.17, QED 0.63
Compound (SMILES): CCC(=O)Nc1cc2c(Nc3ccc4c(ccn4Cc4ccccc4)c3)ncnc2cc1OC
InChIKey: GAYUJMWHGBAPKD-UHFFFAOYSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Irreversible inhibition of GST-tagged ERBB1 (unknown origin) (Met-668 to Ala-1211 residues) expressed in baculovirus infected Sf9 insect cells assessed as reduction in Glu/Tyr copolymer phosphorylation after 6 mins by ELISA
Measurement: IC50 = 21 nM (pChEMBL 7.679999828338623)
Descriptors: MW 451.53, ALogP 5.73, HBA 6, HBD 2, PSA 81.07, QED 0.33
Compound (SMILES): COc1n[nH]c2cc(NC(=O)N[C@@H](c3ccccc3)C3(O)CCCC3)nc(CO)c12
InChIKey: UJFGLYMWAMWCGV-SFHVURJKSA-N
Target: Mitogen-activated protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P28482
Assay: IMAP-FP Assay: Activated ERK2 activity was determined in an IMAP-FP assay (Molecular Devices). Using this assay format, the potency (IC50) of each compound was determined from a 10 point (1:3 serial dilution, 3 uM starting compound concentration) titration curve using the following outlined procedure. To each well of a black Corning 384-well plate (Corning Catalog #3575), 7.5 nL of compound (3333 fold dilution in final assay volume of 25 uL) was dispensed, followed by the addition of 15 uL of kinase buffer (tween containing kinase buffer, Molecular Devices) containing 0.0364 ng/mL (0.833 nM) of phosphorylated active hERK2 enzyme. Following a 15 minute compound & enzyme incubation, each reaction was initiated by the addition of 10 uL kinase buffer containing 2.45 uM ERK2 IMAP substrate peptides (2.25 uM-unlabeled IPTTPITTTYFFFK-COOH and 200 nM-labeled IPTTPITTTYFFFK-SFAM (5-carboxyfluorescein)-COOH), and 75 uM ATP.
Measurement: IC50 = 2.9 nM (pChEMBL 8.539999961853027)
Descriptors: MW 411.46, ALogP 2.63, HBA 6, HBD 5, PSA 132.39, QED 0.42
Compound (SMILES): O=C(O)COc1ccc(Cl)cc1[C@@H]1c2scnc2CCN1C(=O)[C@@H]1C[C@H]1c1ccc(F)cc1
InChIKey: VUQURPMCGGZVDF-GSHUGGBRSA-N
Target: Prostaglandin D2 receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Y4
Assay: Radioligand Displacement Assay: Binding assay was performed in a final assay volume of 250 μl. First, 25 μl of test compound, previously diluted in Binding-Buffer (Binding-Buffer: 50 mM Tris-Base, 100 mM NaCl, 1 mM EDTA, 0.1% BSA (protease free), 0.01% NaN3, 10 mM MnCl2, pH 7.0) was placed into each well. After addition of 75 μl Binding-Buffer, 50 μl of the radioligand 3H-PGD2 (at 2.5 nM (220.000 dpm/well) from ANAWA ART0662) was added to each well. Binding assay was started by addition of 100 μl CRTH2 membrane fragments, reaching a final concentration of 20 μg/well. For non-specific binding, PGD2 was added to the reaction mixture to 10 mM final concentration. This assay mix was incubated for 90 minutes at room temperature and then filtered through a GF/C filter 96-well plate which was pre-soaked for 3 hours in 0.5% polyethyleneimine (PEI). The filter-wells were washed three times with ice cold Binding-Buffer. Then, 40 μl of Microscint-40 (Packard) was added to each well and the re
Measurement: IC50 = 5 nM (pChEMBL 8.300000190734863)
Descriptors: MW 486.95, ALogP 4.68, HBA 5, HBD 1, PSA 79.73, QED 0.55
Compound (SMILES): N#Cc1ccc2sc(C(=O)NC3CCN(Cc4ccc(OC5CCNCC5)c(Br)c4)CC3)cc2c1
InChIKey: OHDBOQZHJURPAR-UHFFFAOYSA-N
Target: Urotensin-2 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UKP6
Assay: Displacement of europium-labeled urotensin-2 from human recombinant urotensin-2 receptor expressed in HEK293 cell membranes after 90 mins by filtration based time resolved fluorometric assay
Measurement: IC50 = 25 nM (pChEMBL 7.599999904632568)
Descriptors: MW 553.53, ALogP 5.06, HBA 6, HBD 2, PSA 77.39, QED 0.45
Compound (SMILES): COc1cc(N(CC2CC2)c2ccc(C(F)(F)F)c3cccnc23)cnc1C(=O)C1CC1C(=O)O
InChIKey: PYVIIPWMEAHZFB-UHFFFAOYSA-N
Target: Leukotriene C4 synthase (SINGLE PROTEIN), Homo sapiens
UniProt: Q16873
Assay: In Vitro hLTC4 Synthase Enzyme Assay (Test A): In the assay, LTC4 synthase catalyses the reaction where the substrate LTA4 is converted to LTC4. Recombinant human LTC4 synthase is expressed in Piccia pastoralis and the purified enzyme is dissolved in 25 mM tris-buffer pH 7.8 supplemented with 0.1 mM glutathione (GSH) and stored at −80 °C. In order to obtain IC50-values for the compounds, the following procedure was used: A volume (24 μL) of 0.25 g/mL LTC4 synthase in 75 mM tris (pH ~8.5), 0.5 mM MgCl2 and 3 mM GSH was preincubated for 10 min with 0.5 μL of compound of interest in DMSO at ten different concentrations, typically in the range 10^−9,5-10^−5 M as well as with DMSO only. The buffer is used as background. Runs are performed in duplicates. The enzymatic reaction is initiated by addition of 0.5 μL LTA4 in diglyme (final assay concentration 8 μM). The reaction is stopped after 1 min by addition of double the reaction volume of a stop solution (MeOH:H2O:acetic acid 70:30:1).
Measurement: IC50 = 229 nM (pChEMBL 6.639999866485596)
Descriptors: MW 485.46, ALogP 5.11, HBA 6, HBD 1, PSA 92.62, QED 0.44
Compound (SMILES): Cc1ncc(C)n2nc(CCc3ccc4cc(C#N)c(Cl)cc4n3)nc12
InChIKey: ZSQKRTOPDCDODI-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y233
Assay: Inhibition Assay: A PDE10A assay may for example, be performed as follows: The assay is performed in 60 uL samples containing a fixed amount of the relevant PDE enzyme (sufficient to convert 20-25% of the cyclic nucleotide substrate), a buffer (50 mM HEPES7.6; 10 mM MgCl2; 0.02% Tween20), 0.1 mg/ml BSA, 225 pCi of 3H-labelled cyclic nucleotide substrate, tritium labeled cAMP to a final concentration of 5 nM and varying amounts of inhibitors. Reactions are initiated by addition of the cyclic nucleotide substrate, and reactions are allowed to proceed for one hr at room temperature before being terminated through mixing with 15 uL 8 mg/mL yttrium silicate SPA beads (Amersham). The beads are allowed to settle for one hr in the dark before the plates are counted in a Wallac 1450 Microbeta counter.
Measurement: IC50 = 59 nM (pChEMBL 7.230000019073486)
Descriptors: MW 362.82, ALogP 3.6, HBA 6, HBD 0, PSA 79.76, QED 0.56
Compound (SMILES): Cc1nc(C)c(-c2csc(Nc3ccc(C(=O)N4[C@@H]5CC[C@H]4C[C@@H](CO)C5)cn3)n2)s1
InChIKey: MQKHRONMZSGXQG-ACHSLPGTSA-N
Target: Transient receptor potential cation channel subfamily V member 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9HBA0
Assay: Antagonist activity at human TRPV4 assessed as inhibition of 4alpha-PDD-induced activation
Measurement: IC50 = 41 nM (pChEMBL 7.389999866485596)
Descriptors: MW 455.61, ALogP 4.4, HBA 8, HBD 2, PSA 91.24, QED 0.59
Compound (SMILES): COc1cc(F)cc(-c2nn(C3Cc4ccccc4C3)c3ncnc(N)c23)c1
InChIKey: GQNBIEAGAUZGAD-UHFFFAOYSA-N
Target: Tyrosine-protein kinase BTK (SINGLE PROTEIN), Homo sapiens
UniProt: Q06187
Assay: Inhibition of human BTK using KVEKIGEGTYGVVYK as substrate after 20 mins by [gamma-33P]ATP based assay
Measurement: IC50 = 1970 nM (pChEMBL 5.710000038146973)
Descriptors: MW 375.41, ALogP 3.56, HBA 6, HBD 1, PSA 78.85, QED 0.59
Compound (SMILES): CCN(CC)CCCC(=O)Nc1nc2cc(C3(O)c4ccccc4C(=O)N3Cc3ccccc3)ccc2[nH]1
InChIKey: YRZZQUOMZIVMFU-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition of B-Raf (unknown origin) pre-incubated for 30 mins by luciferase-luciferin-coupled chemiluminescence assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 511.63, ALogP 4.47, HBA 5, HBD 3, PSA 101.56, QED 0.29
Compound (SMILES): CCOc1ncc2ccccc2c1C(=O)N1[C@@H]2CC[C@H]1[C@H](COc1ccccn1)C2
InChIKey: DQXRZOLGSQYVPN-NSHGMRRFSA-N
Target: Orexin/Hypocretin receptor type 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P56718
Assay: Radioligand Binding Assay: Human Embryonic Kidney 293 cells (HEK293) stably expressing rat orexin 1 receptor (Genebank accession number NM_001525) or Chinese ovary cells (CHO) stably expressing human orexin 1 receptor (Genebank accession number NM_001526) were grown to confluency in DMEM (Hyclone, cat #SH30022), 10% FBS, 1x Pen/Strep, 1x sodium pyruvate, 10 mM HEPES, 600 ug/mL G418 and DMEM/F12 (Gibco, Cat #11039), 10% FBS, 1x Pen/Strep, 600 ug/mL G418 media, respectively on 150 cm^2 tissue culture plates, washed with 5 mM EDTA in PBS (HyClone Dulbecco's Phosphate Buffered Saline 1x with Calcium and Magnesium, Cat #SH30264.01, hereafter referred to simply as PBS) and scraped into 50 ml tubes. After centrifugation (2KxG, 5 min at 4° C.), the supernatant was aspirated and the pellets frozen and stored at -80\°C. Cells were resuspended in PBS in the presence of 1 tablet of protease inhibitor cocktail (Roche, Cat. #11836145001) per 50 mL. Each cell pellet from a 15 cm plate was resuspended in 10 mL, stored on ice, and homogenized for 45 sec prior to addition to the reactions. Competition binding experiments in 96 well polypropylene plates were performed using [3H]-SB674042 (Moraveck Corporation, specific activity=35.3 Ci/mmol), diluted to a 10 nM concentration in PBS (4 nM final). Compounds were solubilized in 100% DMSO (Acros Organics, Cat. #61042-1000) and tested over a range of 7 concentrations (from 0.1 nM to 10 uM). The final concentration of DMSO in the reactions is equal to or less than 0.1%. Total and nonspecific binding was determined in the absence and presence of 10 uM almorexant. The total volume of each reaction is 200 uL (20 uL of diluted compounds, 80 uL of [3H]-(1-(5-(2-fluoro-phenyl)-2-methyl-thiazol-4-yl)-1-((S)-2-(5-phenyl-(1,3,4)oxadiazol-2-ylmethyl)-pyrrolidin-1-yl)-methanone) diluted in PBS and 100 uL of the cell suspension). Reactions were run for 60 min at room temperature and terminated by filtration through GF/C filter plates (PerkinElmer, Cat. #6005174) presoaked in 0.3% polyethylenimine using the cell harvester (PerkinElmer Filtermate). The plates were washed 3 times by aspirating 30 ml PBS through the plates. Plates were dried in 55° C. oven for 60 min, scintillation fluid was added, and the radioactivity was counted on a Topcount (Packard).
Measurement: Ki = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 403.48, ALogP 4.1, HBA 5, HBD 0, PSA 64.55, QED 0.62
Compound (SMILES): CC1(C)CCN(c2ccc(C#Cc3ccccc3)nn2)C(=O)O1
InChIKey: BSWDMNSNGGPJSQ-UHFFFAOYSA-N
Target: Metabotropic glutamate receptor 5 (SINGLE PROTEIN), Homo sapiens
UniProt: P41594
Assay: Intracellular Ca2+ Mobilization Assay: A monoclonal HEK-293 cell line stably transfected with a cDNA encoding for the human mGlu5a receptor was generated; for the work with mGlu5 Positive Allosteric Modulators (PAMs), a cell line with low receptor expression levels and low constitutive receptor activity was selected to allow the differentiation of agonistic versus PAM activity. Cells were cultured according to standard protocols (Freshney, 2000) in Dulbecco's Modified Eagle Medium with high glucose supplemented with 1 mM glutamine, 10% (vol/vol) heat-inactivated bovine calf serum, Penicillin/Streptomycin, 50 μg/ml hygromycin and 15 μg/ml blasticidin (all cell culture reagents and antibiotics from Invitrogen, Basel, Switzerland). bout 24 hrs before an experiment, 5×104 cells/well were seeded in poly-D-lysine coated, black/clear-bottomed 96-well plates. The cells were loaded with 2.5 μM Fluo-4AM in loading buffer (1×HBSS, 20 mM HEPES) for 1 hr at 37 °C. and washed five times with loading buffer. The cells were transferred into a Functional Drug Screening System 7000 (Hamamatsu, Paris, France), and 11 half logarithmic serial dilutions of test compound at 37 °C. were added and the cells were incubated for 10-30 min. with on-line recording of fluorescence. Following this pre-incubation step, the agonist L-glutamate was added to the cells at a concentration corresponding to EC20 (typically around 80 μM) with on-line recording of fluorescence; in order to account for day-to-day variations in the responsiveness of cells, the EC20 of glutamate was determined immediately ahead of each experiment by recording of a full dose-response curve of glutamate. Responses were measured as peak increase in fluorescence minus basal (i.e. fluorescence without addition of L-glutamate), normalized to the maximal stimulatory effect obtained with saturating concentrations of L-glutamate.
Measurement: EC50 = 14 nM (pChEMBL 7.849999904632568)
Descriptors: MW 307.35, ALogP 3, HBA 4, HBD 0, PSA 55.32, QED 0.76
Compound (SMILES): O=c1cc(NC2CCN(S(=O)(=O)C(F)(F)F)CC2)c2cc(C(c3ccc(Cl)cc3)c3ccc(Cl)cc3)cc(NC3CCN(S(=O)(=O)C(F)(F)F)CC3)c2[nH]1
InChIKey: RKRCBMVEJROYQW-UHFFFAOYSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 840.7, ALogP 7.47, HBA 7, HBD 3, PSA 131.68, QED 0.12
Compound (SMILES): NCCCC[C@H](NS(=O)(=O)N[C@@H](CC1CCCCC1)C(=O)N[C@@H]1C[C@H]2CC[C@H]1C2)C(=O)O
InChIKey: PZCYVEWXVUJWOV-FSGKZVOOSA-N
Target: Carboxypeptidase B2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q96IY4
Assay: Inhibition of activated human plasma TAFI incubated for 15 mins by chromogenic assay
Measurement: IC50 = 330 nM (pChEMBL 6.480000019073486)
Descriptors: MW 472.65, ALogP 1.64, HBA 5, HBD 5, PSA 150.62, QED 0.26
Compound (SMILES): Cc1nsc(-c2nnc3n2CCN(C(=O)c2ccc(F)cc2)C3C)n1
InChIKey: PPSNFPASKFYPMN-UHFFFAOYSA-N
Target: Neuromedin-K receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P29371
Assay: Functional Assay: Chinese Hamster Ovary recombinant cells expressing the human NK-3 receptor and a construct that encodes the photoprotein apoaequorin were used for this assay. The antagonist activity of compounds of the invention is measured following pre-incubation (3 minutes) of the compound (at various concentrations) with the cells, followed by addition of the reference agonist (NKA) at a final concentration equivalent to the EC80 (3 nM) and recording of emitted light (FDSS 6000 Hamamatsu) over the subsequent 90-second period. The intensity of the emitted light is integrated using the reader software. Compound antagonist activity is measured based on the concentration-dependent inhibition of the luminescence response to the addition of Neurokinin A.
Measurement: IC50 = 57 nM (pChEMBL 7.239999771118164)
Descriptors: MW 358.4, ALogP 2.46, HBA 7, HBD 0, PSA 76.8, QED 0.7
Compound (SMILES): CN1CC(Cn2cc(-c3ccc(F)c(C(F)(F)F)c3)nc2C2CCN(c3ncnc(N)c3C(N)=O)CC2)C1
InChIKey: GFWUCLLYVKRKSO-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 = 4.9 nM (pChEMBL 8.3100004196167)
Descriptors: MW 532.55, ALogP 3.12, HBA 8, HBD 2, PSA 119.19, QED 0.47
Compound (SMILES): CC[C@@]1(c2ccccc2)NC(=O)N(Cc2ccccc2)C1=O
InChIKey: ZMZDHUHMXXALFX-SFHVURJKSA-N
Target: Cytochrome P450 2C19 (SINGLE PROTEIN), Homo sapiens
UniProt: P33261
Assay: Inhibition of CYP2C19 in human liver microsomes using S-mephenytoin as substrate preincubated for 10 mins followed by NADPH addition measured after 10 mins by LC/MS/MS method
Measurement: IC50 = 138 nM (pChEMBL 6.860000133514404)
Descriptors: MW 294.35, ALogP 3.04, HBA 2, HBD 1, PSA 49.41, QED 0.88
Compound (SMILES): O=C(NO)c1ccc(CNC(=O)c2nc(-c3nccs3)sc2C2CC2)cc1
InChIKey: HPAOMQPZUJIWDB-UHFFFAOYSA-N
Target: Histone deacetylase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13547
Assay: Enzyme Assay: Enzyme activity of HDAC1,3 is determined using the substrate Ac-Lys-Tyr-Lys(Ac)-AMC while the enzyme activity of HDAC6 is assayed using the substrate Boc-lys(Ac)-AMC. The reaction is carried out in flat-bottom, 96-well, or 384-well microplates. After the substrate was deacetylated by HDACs, the product AMC which was obtained from hydrolysis by trypsin can generate fluorescence signal. Measurement is taken in using an multilabe plate reader a 355 nm excitation filter and a 460 nm emission filter. The initial rate of fluorescence should accurately reflect the rate of product formation and enzyme activity. Sample processing: The sample was dissolved in DMSO and kept at a low temperature. DMSO in the final system is limited under a low concentration which won't affect the enzyme activity.
Measurement: IC50 = 42 nM (pChEMBL 7.380000114440918)
Descriptors: MW 400.49, ALogP 3.19, HBA 7, HBD 3, PSA 104.21, QED 0.44
Compound (SMILES): Cc1ccc(-c2ccc(N3CCc4cccc(C(=O)Nc5nc6ccccc6s5)c4C3)nc2C(=O)O)cc1Oc1ccccc1
InChIKey: PALSTDWPMINOGA-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q07817
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The measurement of competition of compounds of Formula (I) with F-Bak for a Bcl-2 family protein (Bcl-xL) binding site using a Time Resolved Fluorescence Resonance Energy Transfer (TR-FRET) binding assay: Test compounds were serially diluted in DMSO starting at 50 M (2x starting concentration; 10% DMSO) and 10 Î¼L transferred into a 384-well plate. Then 10 Î¼L of a protein/probe/antibody mix is added to each well at final concentrations listed in Table 1. Protein: GST-Bcl-xL; 1(nM) Probe: F-Bak (GQVGRQLAIIGDK (6-FAM)INR-amide) SEQ ID NO: 1; 100(nM) Antibody: Tb-anti-GST; 1(nM). The samples are then mixed on a shaker for 1 minute then incubated for an additional 2 hours at room temperature. For each assay plate, a probe/antibody and protein/antibody/probe mixture were included as a negative and a positive control, respectively. Fluorescence was measured on the ENVISION plate reader (Perkin Elmer) using a 340/35 nm excitation filter and 520/525 (F-Bak) and 495/510 nm (Tb-labeled anti-his antibody) emission filters. Dissociation constants (Ki) were determined using Wang's equation (Wang, Z.X. An 20 exact mathematical expression for describing competitive binding of two different ligands to protein molecule. FEBS Lett. 1995 360:111-114). The TR-FRET assay can be performed in the presence of varying concentrations of human serum (HS) or fetal bovine serum (FBS).
Measurement: Ki = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 612.71, ALogP 7.97, HBA 7, HBD 2, PSA 104.65, QED 0.19
Compound (SMILES): CC(C)CC1(Cn2cc(-c3ccc(N4CCc5cccc(C(=O)Nc6nc7ccccc7s6)c5C4)nc3C(=O)O)cn2)CCCCC1
InChIKey: MVQZRHWCVRNVCZ-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q07817
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The measurement of competition of compounds of Formula (I) with F-Bak for a Bcl-2 family protein (Bcl-xL) binding site using a Time Resolved Fluorescence Resonance Energy Transfer (TR-FRET) binding assay: Test compounds were serially diluted in DMSO starting at 50 M (2x starting concentration; 10% DMSO) and 10 Î¼L transferred into a 384-well plate. Then 10 Î¼L of a protein/probe/antibody mix is added to each well at final concentrations listed in Table 1. Protein: GST-Bcl-xL; 1(nM) Probe: F-Bak (GQVGRQLAIIGDK (6-FAM)INR-amide) SEQ ID NO: 1; 100(nM) Antibody: Tb-anti-GST; 1(nM). The samples are then mixed on a shaker for 1 minute then incubated for an additional 2 hours at room temperature. For each assay plate, a probe/antibody and protein/antibody/probe mixture were included as a negative and a positive control, respectively. Fluorescence was measured on the ENVISION plate reader (Perkin Elmer) using a 340/35 nm excitation filter and 520/525 (F-Bak) and 495/510 nm (Tb-labeled anti-his antibody) emission filters. Dissociation constants (Ki) were determined using Wang's equation (Wang, Z.X. An 20 exact mathematical expression for describing competitive binding of two different ligands to protein molecule. FEBS Lett. 1995 360:111-114). The TR-FRET assay can be performed in the presence of varying concentrations of human serum (HS) or fetal bovine serum (FBS).
Measurement: Ki = 4 nM (pChEMBL 8.399999618530273)
Descriptors: MW 648.83, ALogP 8.06, HBA 8, HBD 2, PSA 113.24, QED 0.17
Compound (SMILES): Cc1cccc(-c2ccnc(C(=O)O)n2)c1
InChIKey: PDQHHIFSKQKJBL-UHFFFAOYSA-N
Target: Kynurenine 3-monooxygenase (SINGLE PROTEIN), Homo sapiens
UniProt: O15229
Assay: Inhibition of human KMO
Measurement: IC50 = 17 nM (pChEMBL 7.769999980926514)
Descriptors: MW 214.22, ALogP 2.15, HBA 3, HBD 1, PSA 63.08, QED 0.83
Compound (SMILES): Cc1c(C(=O)Nc2c(C)n(C)n(C3CCCCC3)c2=O)noc1/C=C\C(C)(C)C
InChIKey: DXVFLJBDMVFCLI-SEYXRHQNSA-N
Target: E3 ubiquitin-protein ligase SMURF1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9HCE7
Assay: Inhibition of SMURF1 (unknown origin) in presence of E1/E2/Ub mix measured after 45 mins by europium-labeled streptavidin based TR-FRET assay
Measurement: IC50 = 46 nM (pChEMBL 7.340000152587891)
Descriptors: MW 400.52, ALogP 4.61, HBA 6, HBD 1, PSA 82.06, QED 0.81
Compound (SMILES): C[C@H](c1nc2c(cnn2C2CCCC2)c(=O)[nH]1)N1CC(Cc2ccccc2)C1
InChIKey: BADVCPMBMBUEQL-OAHLLOKOSA-N
Target: High affinity cGMP-specific 3',5'-cyclic phosphodiesterase 9A (SINGLE PROTEIN), Homo sapiens
UniProt: O76083
Assay: Inhibition of PDE9A (unknown origin) using [3H]-cGMP as substrate after 30 mins by microbeta counting method
Measurement: IC50 = 34.8 nM (pChEMBL 7.460000038146973)
Descriptors: MW 377.49, ALogP 3.47, HBA 5, HBD 1, PSA 66.81, QED 0.74
Compound (SMILES): Cn1ccc2cc(OCCCN3CCN(c4cccc5sccc45)CC3)ccc2c1=O
InChIKey: FQLPASRHPPCIMN-UHFFFAOYSA-N
Target: D(2) dopamine receptor (SINGLE PROTEIN), Rattus norvegicus
UniProt: P61169
Assay: Dopamine D2 Receptor Binding Assay: The assay was performed according to the method by Kohler et al. (Kohler C, Hall H, Ogren S O and Gawell L, Specific in vitro and in vivo binding of 3H-raclopride. A potent substituted benzamide drug with high affinity for dopamine D-2 receptors in the rat brain. Biochem. Pharmacol., 1985; 34: 2251-2259). The binding assay was performed using 40 ul of the membrane specimen, 20 ul of [3H]-raclopride (final concentration 1 to 2 nM), 20 ul of a test drug and 50 mM Tris-hydrochloric acid buffer (containing 120 mM NaCl, 5 mM KCl, 2 mM CaCl2, 1 mM MgCl2, pH 7.4) so that the total amount was 200 ul (final dimethylsulfoxide concentration 1%). The reaction was performed at room temperature for 1 hour and terminated by conducting suction filtration with a cell harvester on a glass fiber filter plate. The filter plate made of glass fiber was washed with 50 mM Tris-hydrochloric acid buffer (pH 7.4), and after dried, a microplate liquid scintillation cocktail was added.
Measurement: Ki = 1.9 nM (pChEMBL 8.720000267028809)
Descriptors: MW 433.58, ALogP 4.34, HBA 6, HBD 0, PSA 37.71, QED 0.42
Compound (SMILES): Cc1cnc(Nc2cnn(C)c2)nc1NC1CC2CN(C(=O)CC#N)CC2C1
InChIKey: FXYCRVMSUHVPHV-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: Kinase Assay: JAK1 (h) is incubated with 20 mM Tris/HCl pH 7.5, 0.2 mM EDTA, 500 μM GEEPLYWSFPAKKK, 10 mM MgAcetate and [γ-33P-ATP](specific activity approx. 500 cpm/pmol, concentration as required). The reaction is initiated by the addition of the MgATP mix. After incubation for 40 minutes at room temperature, the reaction is stopped by the addition of 3% phosphoric acid solution. 10 μL of the reaction is then spotted onto a P30 filtermat and washed three times for 5 minutes in 75 mM phosphoric acid and once in methanol prior to drying and scintillation counting.
Measurement: IC50 = 5 nM (pChEMBL 8.300000190734863)
Descriptors: MW 380.46, ALogP 1.82, HBA 8, HBD 2, PSA 111.76, QED 0.81
Compound (SMILES): O=C(N[C@@H](c1ccc(Cl)c(F)c1)c1ccno1)N1CCc2cnc(NC3CCOCC3)nc2C1
InChIKey: IZXDFTLUYDUMQF-NRFANRHFSA-N
Target: Mitogen-activated protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P28482
Assay: Enzymatic Assay: Compounds were tested in an enzymatic assay using human ERK-2 (Mitogen Activated Kinase 1), recombinantly expressed as an n-terminal 6-His fusion protein in E. coli and corresponding to aa 8-360. The substrate used was the fluorescent Omnia peptide S/T17 (Invitrogen of Carlsbad, Calif.; Cat. KNZ1171C). Test compounds were diluted in DMSO in 3-fold serial dilutions at 100x final concentrations. In addition to compound, the assay contained 50 mM HEPES [pH 7.3], 10 mM MgCl2, 1 mM DTT, 0.005% Triton-X100, 5 nM ERK-2 enzyme, 6.25 uM S/T17 peptide substrate and 25 uM ATP (corresponding to the observed Km) for a total reaction volume of 25 uL. The assay was run at ambient temperature in a white 384-well polypropylene plate (Nunc, Inc of Naperville, Ill.; Cat. 267462) collecting data every 50 seconds for approximately 30 minutes on an Envision plate reader (PerkinElmer, Inc. of Waltham, Mass.); Excitation 340 nm/Emission 495 nm.
Measurement: IC50 = 4 nM (pChEMBL 8.399999618530273)
Descriptors: MW 486.94, ALogP 3.71, HBA 7, HBD 2, PSA 105.41, QED 0.57
Compound (SMILES): COc1ccc(Cl)c(N2CCC3(CCC(CC(=O)O)CC3)CC2)c1
InChIKey: YIXZOCHHRTUZKL-UHFFFAOYSA-N
Target: Free fatty acid receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5NUL3
Assay: Agonist activity at ARMS2-PK2 -tagged human GPR120 short isoform expressed in CHOK1 cells assessed as induction of beta-arrestin2 recruitment after 90 mins by luminescence assay
Measurement: EC50 = 2500 nM (pChEMBL 5.599999904632568)
Descriptors: MW 351.87, ALogP 4.6, HBA 3, HBD 1, PSA 49.77, QED 0.86
Compound (SMILES): COc1c(Cl)cccc1-c1cc(F)cc(C(CC(=O)O)c2ncco2)c1
InChIKey: YWNUALHNZDLMNL-UHFFFAOYSA-N
Target: Lysosomal protective protein (SINGLE PROTEIN), Homo sapiens
UniProt: P10619
Assay: Inhibition Assay: Recombinant human cathepsin A (residues 29-480, with a C-terminal 10-His tag; R&D Systems, #1049-SE) was proteolytically activated with recombinant human cathepsin L (R&D Systems, #952-CY). Briefly, cathepsin A was incubated at 10 ug/ml with cathepsin L at 1 ug/ml in activation buffer (25 mM 2-(morpholin-4-yl)-ethanesulfonic acid (MES), pH 6.0, containing 5 mM dithiothreitol (DTT)) for 15 min at 37° C. Cathepsin L activity was then stopped by the addition of the cysteine protease inhibitor E-64 (N-(trans-epoxysuccinyl)-L-leucine-4-guanidinobutylamide; Sigma-Aldrich, # E3132; dissolved in activation buffer/DMSO) to a final concentration of 10 uM.The activated cathepsin A was diluted in assay buffer (25 mM MES, pH 5.5, containing 5 mM DTT) and mixed with the test compound (dissolved in assay buffer containing (v/v) 3% DMSO) or, in the control experiments, with the vehicle in a multiple assay plate. After incubation for 15 min at room temperature.
Measurement: IC50 = 815 nM (pChEMBL 6.090000152587891)
Descriptors: MW 375.78, ALogP 4.75, HBA 4, HBD 1, PSA 72.56, QED 0.67
Compound (SMILES): O=C(Nc1ccccc1)N1CCN(c2ccnc3cc(Cl)ccc23)CC1
InChIKey: ICASXVHIXTXGMF-UHFFFAOYSA-N
Target: DNA gyrase subunit B (SINGLE PROTEIN), Mycobacterium smegmatis (strain ATCC 700084 / mc(2)155)
UniProt: A0QNE0
Assay: Inhibition of Mycobacterium smegmatis 155 6His-tagged DNA gyrase B catalytic domain ATPase activity expressed in Escherichia coli BL21 (DE3) pLysS cells assessed as inorganic phosphate release after 120 mins in presence of ATP by malachite green dye based assay
Measurement: IC50 = 7600 nM (pChEMBL 5.119999885559082)
Descriptors: MW 366.85, ALogP 4.24, HBA 3, HBD 1, PSA 48.47, QED 0.74
Compound (SMILES): O=C(CCOc1ccccc1)N1CCN(C(=O)n2sc3ncccc3c2=O)CC1
InChIKey: AGXGWHOSTWFVQH-UHFFFAOYSA-N
Target: Protein-glutamine gamma-glutamyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P21980
Assay: TGM2 Inhibitory Activity Assay: 2 ul of a solution of different concentrations of the test compound in dimethyl sulfoxide was added to 28 ul of buffer (50 mM TRIS, 100 mM NaCl, 10 mM CaCl2, pH 7.5) and 10 ul of a TGM2 solution to give a TMG2 test concentration of 25 ug/ml, and the mixture incubated for 15 minutes at room temperature in a 96 half-well microtiter plate. The enzyme reaction was started by the addition of 10 ul of substrate solution containing H-Abz-APE(CAD-DNP)QEA-OH and glycine methyl ester hydrochloride in buffer to give test concentrations of H-Abz-APE(CAD-DNP)QEA-OH of 50 uM and of glycine methyl ester hydrochloride of 5 mM. The time course of the reaction was monitored with excitation at 318 nm and measurement at emission wavelength 418 nm in a microtiter plate reader (SpectraMax M5, Molecular Devices) over 15 minutes. Rates were calculated from the linear part of the curve (generally between 5 and 10 minutes). IC50 values were calculated from the means (duplicates) of a dilution series of the com
Measurement: IC50 = 110 nM (pChEMBL 6.960000038146973)
Descriptors: MW 412.47, ALogP 2.04, HBA 7, HBD 0, PSA 84.74, QED 0.65
Compound (SMILES): O=C(O)Cc1ccc(OC2CC2)c(-c2ccc(F)c3c2CN(C(=O)OCc2ccccc2)CC3)c1
InChIKey: LYUOPXOHOYOKSW-UHFFFAOYSA-N
Target: Prostaglandin D2 receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Y4
Assay: Radioligand Displacement Assay: Binding assay was performed in a final assay volume of 250 ul. First, 25 ul of test compound, previously diluted in Binding-Buffer (Binding-Buffer: 50 mM Tris-Base, 100 mM NaCl, 1 mM EDTA, 0.1% BSA (protease free), 0.01% NaN3, 10 mM MnCl2, pH 7.0) was placed into each well. After addition of 75 ul Binding-Buffer, 50 ul of the radioligand 3H-PGD2 (at 2.5 nM (220.000 dpm/well) from ANAWA ART0662) was added to each well. Binding assay was started by addition of 100 ul CRTH2 membrane fragments, reaching a final concentration of 20 ug/well. For non-specific binding, PGD2 was added to the reaction mixture to 10 mM final concentration. This assay mix was incubated for 90 minutes at room temperature and then filtered through a GF/C filter 96-well plate which was pre-soaked for 3 hours in 0.5% polyethyleneimine (PEI). The filter-wells were washed three times with ice cold Binding-Buffer. Then, 40 ul of Microscint-40 (Packard) was added to each well and the retained radioactivity quantified in a Topcount (Packard).
Measurement: IC50 = 14.9 nM (pChEMBL 7.829999923706055)
Descriptors: MW 475.52, ALogP 5.36, HBA 4, HBD 1, PSA 76.07, QED 0.5
Compound (SMILES): Cc1cc(-c2nc3cc(S(=O)(=O)N(C)C)cnc3n2Cc2ccccc2)cn(C)c1=O
InChIKey: NZTUGGCQTGSXPM-UHFFFAOYSA-N
Target: Bromodomain-containing protein 4 (SINGLE PROTEIN), Homo sapiens
UniProt: O60885
Assay: AlphaScreen Assay: Compounds are diluted in serial dilution 1:5 in assay buffer from 10 mM stock in DMSO (100 μM start concentration) in white OptiPlate-384 (PerkinElmer). A mix consisting of 15 nM GST-BRD4-BD1 protein (aa 44-168) or 150 nM GST-BRD4-BD2 (aa 333-460) and 15 nM biotinylated Acetyl-Histone H4 (LysS, 8, 12, 16) peptide is prepared in assay buffer (50 mM HEPES pH=7.4; 25 mM NaCl; 0.05% Tween 20; 0.1% bovine serum albumin (BSA); 10 mM dithiothreitol (DTT)). 6 μl of the mix is added to the compound dilutions. Subsequently, 6 μl of premixed AlphaLISA Glutathione Acceptor Beads and AlphaScreen Streptavidin Donor Beads from PerkinElmer (in assay buffer at a concentration of 10 μg/ml each) are added and the samples are incubated for 30 min at RT in the dark (shaking 300 rpm). Afterwards, the signal is measured in a PerkinElmer Envision HTS Multilabel Reader using the AlphaScreen protocol from PerkinElmer.
Measurement: IC50 = 119 nM (pChEMBL 6.920000076293945)
Descriptors: MW 437.53, ALogP 2.4, HBA 7, HBD 0, PSA 90.09, QED 0.48
Compound (SMILES): CN(C)c1ncc(S(=O)(=O)c2ccc(CNC(=O)N3Cc4ccncc4C3)nc2)s1
InChIKey: PJFVLPUONQTLGX-UHFFFAOYSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Biochemical Inhibition Assay: The NAMPT Enzymatic Reaction. The NAMPT enzymatic reactions were carried out in Buffer A (50 mM Hepes pH 7.5, 50 mM NaCl, 5 mM MgCl2, and 1 mM THP) in 96-well V-bottom plates. The compound titrations were performed in a separate dilution plate by serially diluting the compounds in DMSO to make a 100x stock. Buffer A (89 uL) containing 33 nM of NAMPT protein was added to 1 uL of 100x compound plate containing controls (e.g. DMSO or blank). The compound and enzyme mixture was incubated for 15 min at rt, then 10 uL of 10x substrate and co-factors in Buffer A were added to the test well to make a final concentration of 1 uM NAM, 100 pM 5-Phospho-D-ribose 1-diphosphate (PRPP), and 2.5 mM Adenosine 5'-triphosphate (ATP). The reaction was allowed to proceed for 30 min at rt, then was quenched with the addition of 11 uL of a solution of formic acid and L-Cystathionine to make a final concentration of 1% formic acid and 10 uM L-Cystathionine. Background and signal strength was determined by addition (or non-addition) of a serial dilution of NMN to a pre-quenched enzyme and cofactor mix.
Measurement: IC50 = 6.6 nM (pChEMBL 8.180000305175781)
Descriptors: MW 444.54, ALogP 2.06, HBA 8, HBD 1, PSA 108.39, QED 0.64
Compound (SMILES): O=C(Cc1cccc(Cl)c1)Nc1cccc(CN2CCC(C(=O)NC3CCCCC3)CC2)c1
InChIKey: XMNRYCLMXUYTCH-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 10 nM (pChEMBL 8)
Descriptors: MW 468.04, ALogP 5.18, HBA 3, HBD 2, PSA 61.44, QED 0.59
Compound (SMILES): CCC(=O)NC1CCc2c(-c3cc4c(cc3F)N(C)C(=O)CC4)cncc21
InChIKey: RXJOFZNMHGDABT-UHFFFAOYSA-N
Target: Cytochrome P450 11B2, mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: P19099
Assay: Cellular Enzyme Assay: The expression plasmids contains the ORF for either human/cyno CYP11B1 or CYP11B2 under the control of a suitable promoter (CMV-promoter) and a suitable resistance marker (neomycin). Using standard techniques the expression plasmid is transfected into G-402 cells and these cells are then selected for expressing the given resistance markers. Individual cell-clones are then selected and assessed for displaying the desired enzymatic activity using 11-Deoxycorticosterone (Cyp11B2) or 11-Deoxycortisol (Cyp11B1) as a substrate. G-402 cells expressing CYP11 constructs were established as described above and maintained in McCoy's 5a Medium Modified, ATCC Catalog No. 30-2007 containing 10% FCS and 400 μg/ml G418 (Geneticin) at 37° C. under an atmosphere of 5% CO2/95% air. Cellular enzyme assays were performed in DMEM/F12 medium containing 2.5% charcoal treated FCS and appropriate concentration of substrate (0.3-10 μM 11-Deoxycorticosterone, 11-Deoxycortisol or Corticosterone). For assaying enzymatic activity, cells were plated onto 96 well plates and incubated for 16 h. An aliquot of the supernatant is then transferred and analyzed for the concentration of the expected product (Aldosterone for CYP11B2; Cortisol for CYP11B1). The concentrations of these steroids can be determined using HTRF assays from CisBio analyzing either Aldosterone or Cortisol.
Measurement: EC50 = 10.2 nM (pChEMBL 7.989999771118164)
Descriptors: MW 367.42, ALogP 3.31, HBA 3, HBD 1, PSA 62.3, QED 0.91