lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CCOC(=O)COc1cccc2ccc(Cc3cncc4cc(OC)c(OC)cc34)nc12
InChIKey: RDBLVTMTIJBORU-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y233
Assay: Diaxonhit Phosphodiesterase Assay: The phosphodiesterase assay was developed using the LANCE cAMP kit (PerkinElmer). The assay buffer contained HBSS with 5 mM HEPES, 0.1% BSA, and 1.5 mM MgCl2, pH 7.4. PDE10A (BPS Bioscience) was used at 200 pg/well (with a specific activity of 3200 pmole/min/ug with assay conditions: 10 mM Tris-HCl, pH 7.4, 10 mM MgCl2, 1 mM MnCl2, 200 uM cAMP, 2.5 kU 5' nucleotidase, 37° C., 20 min) and PDE4D3 (BPS Bioscience) was used at 100 pg/well (with a specific activity of 32713 pmole/min/ug with assay conditions: 10 mM Tris-HCl, pH 7.4, 10 mM MgCl2, 1 mM MnCl2, 200 uM cAMP, 2.5 kU 5' nucleotidase, 37° C., 20 min). The Biotin-cAMP tracer, supplied in 10 mmol/L Tris-HCl buffered (pH 8.0) salt solution with 1 mmol/L ethylenediaminetetraacetic acid (EDTA), 0.1% bovine serum albumin (BSA), and 0.05% sodium azide, is used at a dilution of 1/375. The assay detection mixture contained the LANCE Eu-W8044 labeled streptavidin 1/2250 (supplied in 50 mmol/L Tris-HCl buffer).
Measurement: IC50 = 51 nM (pChEMBL 7.289999961853027)
Descriptors: MW 432.48, ALogP 4.33, HBA 7, HBD 0, PSA 79.77, QED 0.38
Compound (SMILES): CC(C)(C)CN1Cc2[nH]nc(C(=O)N3CCC(c4ccc(F)c(F)c4C(F)(F)F)CC3)c2C1
InChIKey: FFWLYIPCGDWQBE-UHFFFAOYSA-N
Target: Retinol-binding protein 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P02753
Assay: Scintillation Proximity Binding Assay: Untagged human RBP4 purified from urine of tubular proteinuria patients was purchased from Fitzgerald Industries International. It was biotinylated using the EZ-Link Sulfo-NHS-LC-Biotinylation kit from Pierce following the manufacturer's recommendations. Binding experiments were performed in 96-well plates (OptiPlate, PerkinElmer) in a final assay volume of 100 ul per well in SPA buffer (IX PBS, pH 7.4, 1 mM EDTA, 0.1% BSA, 0.5% CHAPS). The reaction mix contained 10 nM 3H-Retinol (48.7 Ci/mmol; PerkinElmer), 0.3 mg/well Streptavidin-PVT beads, 50 nM biotinylated RBP4 and a test compound. Nonspecific binding was determined in the presence of 20 uM of unlabeled retinol. The reaction mix was assembled in the dark under dim red light. The plates were sealed with clear tape (TopSeal-A: 96-well microplate, PerkinElmer), wrapped in the aluminum foil, and allowed to equilibrate 6 hours at room temperature followed by overnight incubation at +4° C. Radiocounts were measured using a TopCount NXT counter (Packard Instrument Company).
Measurement: IC50 = 22.48 nM (pChEMBL 7.650000095367432)
Descriptors: MW 470.49, ALogP 5.09, HBA 3, HBD 1, PSA 52.23, QED 0.64
Compound (SMILES): CSc1ncc2c(n1)CCN(c1cc(C(=O)Nc3cccc(C(F)(F)F)c3)ccc1C)C2
InChIKey: VRLCFROVLGMZQU-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: VEGFR2 Kinase Assay: Biochemical KDR kinase assays were performed in 96 well microliter plates that were coated overnight with 75 μg/well of poly-Glu-Tyr (4:1) in 10 mM Phosphate Buffered Saline (PBS), pH 7.4. The coated plates were washed with 2 mls per well PBS+0.05% Tween-20 (PBS-T), blocked by incubation with PBS containing 1% BSA, then washed with 2 mls per well PBS-T prior to starting the reaction. Reactions were carried out in 100 μL reaction volumes containing 2.7 Î¼M ATP in kinase buffer (50 mM Hepes buffer pH 7.4, 20 mM MgCl2, 0.1 mM MnCl2and 0.2 mM Na3VO4). Test compounds were reconstituted in 100% DMSO and added to the reaction to give a final DMSO concentration of 5%. Reactions were initiated by the addition 20 μl per well of kinase buffer containing 200-300 ng purified cytoplasmic domain KDR protein (BPS Bioscience, San Diego, Calif.). Following a 15 minute incubation at 30° C., the reactions were washed 2 mls per well PBS-T. 100 μl of a monoclonal anti-phosphotyrosine antibody-peroxidase conjugate diluted 1:10,000 in PBS-T was added to the wells for 30 minutes. Following a 2 mls per well wash with PBS-Tween-20, 100 μl of O-Phenylenediamine Dihydrochloride in phosphate-citrate buffer, containing urea hydrogen peroxide, was added to the wells for 7-10 minutes as a colorimetric substrate for the peroxidase. The reaction was terminated by the addition of 100 μl of 2.5N H2SO4 to each well and read using a microplate ELISA reader set at 492 nm.
Measurement: IC50 = 71 nM (pChEMBL 7.150000095367432)
Descriptors: MW 458.51, ALogP 5.34, HBA 5, HBD 1, PSA 58.12, QED 0.42
Compound (SMILES): CC(C)(NS(=O)(=O)c1ccc(NC2CCCCC2)cc1C(F)(F)F)C1CC1
InChIKey: SAFPVFNYBFBXRR-UHFFFAOYSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
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: Ki = 6000 nM (pChEMBL 5.21999979019165)
Descriptors: MW 404.5, ALogP 4.92, HBA 3, HBD 2, PSA 58.2, QED 0.71
Compound (SMILES): Nc1cccc(N2C(=O)c3ccccc3C2(O)c2ccc3c(c2)NC(=O)CO3)c1
InChIKey: RWJGXHSKKVZYLI-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 387.4, ALogP 2.45, HBA 5, HBD 3, PSA 104.89, QED 0.59
Compound (SMILES): CNC(=O)c1cccc(Nc2ncnc(N3CCC(c4nc(-c5ccccc5)cs4)CC3)n2)c1
InChIKey: APSHEFAEYJWLEY-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Cell-Based Assay: Compounds were tested on human Nav1.7 (hNav1.7) expressed stably in 293 cells, which have no endogenous sodium channels. Two forms of whole-cell patch-clamp assays were used. In both, peak current through Nav1.7 was evoked by a depolarizing voltage pulse, and currents were monitored as a function of time in response to exposure to several different concentrations of drug. Automated electrophysiology uses the PatchXpress 7000A system. Up to sixteen cells at a time were voltage-clamped in parallel, and the steady-state voltage dependence of inactivation was determined experimentally for each cell. For each cell, fractional inactivation was set at 20%, and peak current through Nav1.7 as a function of time was measured in response to increasing concentrations of test compound. Manual electrophysiology refers to conventional whole-cell patch-clamping (Hamill et al., 1981). After formation of the whole-cell clamp, cells were lifted off the bottom of the dish with the patch pipette and positioned directly in front of an array of glass microperfusion tubes each with internal diameter about 250 microns, with individual compound concentrations flowing from each tube. Current as a function of time in response to increasing drug concentrations was measured with Nav1.7 fully in the resting state (holding voltage set to −140 mV) and on the same cells with the holding voltage set to produce about 20% inactivation. Compounds were not applied for a prespecified time, but for as long as required to achieve a stable level of block.
Measurement: IC50 = 380 nM (pChEMBL 6.420000076293945)
Descriptors: MW 471.59, ALogP 4.48, HBA 8, HBD 2, PSA 95.93, QED 0.43
Compound (SMILES): COc1ccc2ccn(CCCCN3CCN(c4cccc5sccc45)CC3)c(=O)c2c1
InChIKey: VNFXFHGUVVNZTJ-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 = 1.2 nM (pChEMBL 8.920000076293945)
Descriptors: MW 447.6, ALogP 4.83, HBA 6, HBD 0, PSA 37.71, QED 0.38
Compound (SMILES): Oc1ccccc1CCCNCc1ccccc1O
InChIKey: WBOAWYQEMHFWQV-UHFFFAOYSA-N
Target: Myeloperoxidase (SINGLE PROTEIN), Homo sapiens
UniProt: P05164
Assay: Inhibition of MPO chlorination activity (unknown origin) assessed as taurine chloramine formation after 5 mins in presence of H2O2/Cl- by HTS method
Measurement: IC50 = 350 nM (pChEMBL 6.460000038146973)
Descriptors: MW 257.33, ALogP 2.82, HBA 3, HBD 3, PSA 52.49, QED 0.7
Compound (SMILES): COc1cc(C(=O)N2CC[C@H]2c2nc(-c3cccc(F)c3OC)no2)c(-n2nccn2)cc1C
InChIKey: HQHRHVVXQYRDCR-KRWDZBQOSA-N
Target: Orexin/Hypocretin receptor type 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O43613
Assay: Intracellular Calcium Assay: 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 μg/mL G418, 100 U/mL penicillin, 100 μg/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 μM 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. Within the Fluorescent Imaging Plate Reader (FLIPR Tetra, Molecular Devices), antagonists are added to the plate in a volume of 10 μL/well, incubated for 120 min and finally 10 μL/well of agonist is added. Fluorescence is measured for each well at 1 second intervals, and the height of each fluorescence peak is compared to the height of the fluorescence peak induced by 3 nM orexin-A with vehicle in place of antagonist.
Measurement: IC50 = 439 nM (pChEMBL 6.360000133514404)
Descriptors: MW 464.46, ALogP 3.37, HBA 9, HBD 0, PSA 108.4, QED 0.43
Compound (SMILES): O=C(O)CC(c1cccc(-c2ccc(OCC3CC3)cc2)c1)c1nccs1
InChIKey: ZVVKMTYXBPZAPY-UHFFFAOYSA-N
Target: Lysosomal protective protein (SINGLE PROTEIN), Homo sapiens
UniProt: P10619
Assay: Inhibition Assay: 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, as substrate then bradykinin carrying an N-terminal® Bodipy FL (4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl) label (JPT Peptide Technologies GmbH; dissolved in assay buffer) was added to the mixture. The final concentration of cathepsin A was 833 ng/ml and the final concentration of labeled bradykinin 2 μM. After incubation for 15 min at room temperature the reaction was stopped by the addition of stop buffer (130 mM 2-(4-(2-hydroxy-ethyl)-piperazin-1-yl)-ethanesulfonic acid, pH 7.4, containing (v/v) 0.013%° Triton X-100, 0.13% Coating Reagent 3 (Caliper Life Sciences), 6.5% DMSO and 20 μM ebelactone B (Sigma, #E0886)).
Measurement: IC50 = 2290 nM (pChEMBL 5.639999866485596)
Descriptors: MW 379.48, ALogP 5.21, HBA 4, HBD 1, PSA 59.42, QED 0.58
Compound (SMILES): Fc1ccc(-c2cc(CNc3nc(Nc4cc(C5CC5)n[nH]4)cc(OCCN4CCOCC4)n3)on2)cc1
InChIKey: VAKLBUXAUZVGEE-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) un-phosphorylated apo-enzyme form expressed in baculovirus based expression system using poly-EY peptide incubated for 2 hrs by luciferase-luciferin coupled chemiluminescent assay
Measurement: IC50 = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 520.57, ALogP 3.94, HBA 10, HBD 3, PSA 126.25, QED 0.27
Compound (SMILES): Cc1c(C(=O)Nc2ccc(Oc3ccnc4cc(OCC(C)(C)O)ccc34)cc2)c(=O)n(-c2ccccc2)n1C
InChIKey: JNVFMMAYSXTMTJ-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Kinase Assay: Met (h) is incubated with 8 mM MOPS pH 7.0, 0.2 mM EDTA, 250 μM KKKSPGEYVNIEFG, 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 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 538.6, ALogP 5.23, HBA 8, HBD 2, PSA 107.61, QED 0.28
Compound (SMILES): Cc1cc(NC(=O)Nc2ccc(Cl)cc2)ccc1S(=O)(=O)Nc1ccccc1S(N)(=O)=O
InChIKey: QSCZKUJMKKBSPL-UHFFFAOYSA-N
Target: Prostaglandin E synthase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9H7Z7
Assay: Biochemical Assay: A solution of test compound was added to a diluted microsome preparation containing human mPGES-1 and pre-incubated for 15 minutes in potassium phosphate buffer pH 6.8 with cofactor glutathione (GSH). Corresponding solutions without test compound were used as positive controls, and corresponding solutions without test compound and without microsomes were used as negative controls. The enzymatic reaction was then started by addition of the substrate PGH2 in an organic solution (dry acetonitrile).The typical reaction conditions of the enzymatic reaction were thus: Test compound: ranging from 60 uM to 0.002 uM, or zero in positive and negative controls; potassium phosphate buffer pH 6.8: 50 mM; GSH: 2.5 mM; mPGES-1-containing microsomes: 2 ug/mL (sample and positive controls) or 0 ug/mL (negative control); PGH2: 10.8 uM; Acetonitrile: 7.7% (v/v); DMSO: 0.6% (v/v). The reaction was stopped after one minute by adding an acidic solution (pH 1.9) of ferric chloride.
Measurement: IC50 = 900 nM (pChEMBL 6.050000190734863)
Descriptors: MW 494.98, ALogP 3.74, HBA 5, HBD 4, PSA 147.46, QED 0.41
Compound (SMILES): CC(=O)c1ccc(OCc2cccc(F)c2)cc1O
InChIKey: UFWGBYKUPYCCBE-UHFFFAOYSA-N
Target: Amine oxidase [flavin-containing] B (SINGLE PROTEIN), Homo sapiens
UniProt: P27338
Assay: Inhibition of recombinant human MAO-B using p-tyramine as substrate assessed as decrease in H2O2 production incubated for 15 mins by fluorimetric method
Measurement: IC50 = 41.38 nM (pChEMBL 7.380000114440918)
Descriptors: MW 260.26, ALogP 3.31, HBA 3, HBD 1, PSA 46.53, QED 0.86
Compound (SMILES): O=C(C1CCS(=O)(=O)CC1)N1CCN(c2ccccn2)CC1c1cccc(F)c1F
InChIKey: PUCVVBOYBJSZKV-UHFFFAOYSA-N
Target: Sodium- and chloride-dependent glycine transporter 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P48067
Assay: Cytostar-T Assay: In detail, SK-N-MC cells are seeded into 96-well Cytostar-T assay plates at a density of 200,000 cells/well and grown for 16-18 hours to confluence in growth medium as recommended by ATCC. Before starting the assay, cells are washed once with HBSS (Hank's buffered salt solution; Sigma, H8264) cont. 5 mM alanine (referred in here as HBSS/Ala) and afterwards the following reagents are added: 1.80 ul/well HBSS/A1a 2. 20 ul/well of HBSS/A1a containing 6x the concentration of compound in 6% DMSO 3. approx. 5-10 min incubation 4. 20 ul/well 3 uM glycine (3H-glycine (Perkin Elmer, NET004001MC, specific activity: 52 Ci/mmol; diluted 1:1 with unlabelled glycine) in HBSS/A1a. In the final assay, glycine concentration is 500 nM (250 nM derived from the 3H-glycine Perkin Elmer, 250 nM unlabelled glycine), DMSO concentration is 1%. The assay plate is immediately after addition of the 3H-glycine placed into a Micro-Beta Counter (Perkin Elmer) and the signal is recorded over 60 min.
Measurement: IC50 = 98 nM (pChEMBL 7.010000228881836)
Descriptors: MW 435.5, ALogP 2.57, HBA 5, HBD 0, PSA 70.58, QED 0.74
Compound (SMILES): CC(C)n1cc(C(=O)c2cncc(NC(=O)Cc3nnn[nH]3)c2)c2cnccc21
InChIKey: KOCPMGCABJFCDA-UHFFFAOYSA-N
Target: High affinity nerve growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P04629
Assay: HTRF Assay: Isolated TRK Enzyme assays use the HTRF KinEASE-TK kit (Cisbio Cat#62TK0PEJ) with recombinant His-tagged cytoplasmic domains of each TRK receptor sourced from Invitrogen (see table below). This activity-assay measures the phosphorylation of tyrosine residues within a substrate from the HTRF kit which has been validated by Cisbio for a variety of tyrosine kinases including the TRK receptors.Assay Details: Invitrogen Amino FAC FAC Assay ReactionTarget Cat# acids enzyme ATP TimeTRKA PV3144 aa 441-4 nM 40 uM 35 min (NTRK1) 796 TRKB PV3616 aa 526-1 nM 1.4 uM 40 min (NTRK2) 838 TRKC PV3617 aa 510- 10 nM 15 uM 30 min (NTRK3) 8250.5 mM stock solutions of test compounds are prepared and serially diluted in 100% DMSO. A standard curve using the compound of Example 135 disclosed in WO2005/116035 of 150 uM is also prepared on each test plate. High percentage effect (HPE) is defined by 150 uM.
Measurement: IC50 = 9800 nM (pChEMBL 5.010000228881836)
Descriptors: MW 390.41, ALogP 1.94, HBA 8, HBD 2, PSA 131.34, QED 0.48
Compound (SMILES): C=CC(=O)Nc1cccc(-c2cnc3[nH]cc(-c4cc(C)c(F)cc4OC)c3c2)c1
InChIKey: ZNSLGAKNFMLWFV-UHFFFAOYSA-N
Target: Tyrosine-protein kinase ITK/TSK (SINGLE PROTEIN), Homo sapiens
UniProt: Q08881
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 > 1000 nM (pChEMBL 6)
Descriptors: MW 401.44, ALogP 5.48, HBA 3, HBD 2, PSA 67.01, QED 0.44
Compound (SMILES): CN1CCN(Cc2ccc(-c3cn4c5nc(ncc35)NCCCCCC(O)CCCC4)cc2)CC1
InChIKey: SDWTWVXMBPUQSC-UHFFFAOYSA-N
Target: Tyrosine-protein kinase Mer (SINGLE PROTEIN), Homo sapiens
UniProt: Q12866
Assay: Inhibition of human MerTK kinase domain (1585 to 3000 residues) expressed in HEK293 cells co-expressing rat EGFR LBD assessed as inhibition of EGF-stimulated MerTK phosphorylation preincubated for 30 mins followed by EGF stimulation for 15 mins by ELISA
Measurement: EC50 = 150 nM (pChEMBL 6.820000171661377)
Descriptors: MW 476.67, ALogP 4.36, HBA 7, HBD 2, PSA 69.45, QED 0.59
Compound (SMILES): N#Cc1ccc2c3c(ccc2c1)O[C@@H](c1cc(F)c(F)cc1F)[C@H](N)C3
InChIKey: IHMRKDWNSLQDOV-QUCCMNQESA-N
Target: Dipeptidyl peptidase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P27487
Assay: Inhibition of DPP4 (unknown origin) expressed in Sf9 cells using Gly-Pro-AMC substrate
Measurement: IC50 = 8.42 nM (pChEMBL 8.069999694824219)
Descriptors: MW 354.33, ALogP 4.13, HBA 3, HBD 1, PSA 59.04, QED 0.67
Compound (SMILES): Cc1c(NC(=O)c2ccc(C(C)(C)O)cc2F)cc(F)cc1-c1ncnc2[nH]c(C3=CCN(C(=O)N(C)C)CC3)cc12
InChIKey: MLKDBPSLDGNZKN-UHFFFAOYSA-N
Target: Tyrosine-protein kinase BTK (SINGLE PROTEIN), Homo sapiens
UniProt: Q06187
Assay: Biochemical Enzyme Assay: The inhibitory activity of the present compounds against Btk was assessed in a biochemical enzyme assay. Assay plates in 384 well format were prepared with 8-point serial dilutions for the test compounds on a Thermo CatX workstation equipped with a Innovadyne Nanodrop Express. The assay plates were prepared by addition of 50 nl per well of compound solution in 90% DMSO. The kinase reactions were started by stepwise addition of 4.5 ul per well of peptide/ATP-solution (4 uM FITC-Ahx-TSELKKVVALYDYMPMNAND-NH2, 164 uM ATP) in kinase buffer (50 mM HEPES, pH 7.5, 1 mM DTT, 0.02% Tween20, 0.02% BSA, 0.6% DMSO, 10 mM beta-glycerophosphate, and 10 uM sodium orthovanadate, 18 mM MgCl2, 1 mM MnCl2) and 4.5 ul per well of enzyme solution (6.4 nM full-length human recombinant BTK) in kinase buffer. Kinase reactions were incubated at 30° C. for 60 minutes and subsequently terminated by addition of 16 ul per well of stop solution (100 mM HEPES pH 7.5, 5% DMSO, 0.1% Caliper coating reagent, 10 mM EDTA, and 0.015% Brij35). Kinase reactions were analyzed on a Caliper LC3000 workstation by separating phosphorylated and unphosphorylated peptides and kinase activities were calculated from the amounts of newly formed phospho-peptide. Inhibition data were calculated by comparison to control reactions without enzyme (100% inhibition) and without inhibitors (0% inhibition).
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 574.63, ALogP 5.46, HBA 5, HBD 3, PSA 114.45, QED 0.29
Compound (SMILES): O=C(O)CNC(=O)COC[C@H]1CC[C@H](COC(=O)N(c2ccccc2)c2ccc(Cl)cc2)CC1
InChIKey: LPELAFLWIGWDFH-WGSAOQKQSA-N
Target: Prostacyclin receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P43119
Assay: Agonist activity at human recombinant prostacyclin receptor expressed in CHO-K1 cells assessed as induction of cAMP accumulation incubated for 1 hr by HTRF assay
Measurement: EC50 = 2.7 nM (pChEMBL 8.569999694824219)
Descriptors: MW 488.97, ALogP 4.64, HBA 5, HBD 2, PSA 105.17, QED 0.5
Compound (SMILES): Cc1cccc(NC(=O)Nc2ccc(-c3cncc4[nH]nc(N)c34)c3ccccc23)c1
InChIKey: JCKMPGSSOFUEAB-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Enzyme-Linked Immunosorbent Assay: Experimental Procedures(1) Kinase reaction substrate Poly(Glu, Tyr, 4:1) was diluted to 20 Î¼g/ml with potassium ion-free PBS, and enzyme labeled plate was coated by the substrate, reacted at 37° C. for 12-16 h, and then the liquid in holes was removed.(2) Enzyme labeled plate was washed with T-PBS for three times, 10 min each time.(3) Enzyme labeled plate was dried at 37° C. in a dryer.(4) The tested samples were added into holes of the coated enzyme labeled plate:The tested samples were firstly formulated to be a 10â¿¿2 M stock solution with DMSO, and then diluted to required concentration with reaction buffer before use, added into the experiment holes to reach corresponding final concentration in a 100 uL reaction system. Meanwhile, positive control holes were created, and compound Sorafenib was added. The rest of the stock solutions was storaged at â¿¿20° C. after subpackage.
Measurement: IC50 = 4.3 nM (pChEMBL 8.369999885559082)
Descriptors: MW 408.47, ALogP 5.31, HBA 4, HBD 4, PSA 108.72, QED 0.32
Compound (SMILES): O=C(c1ccc2cc(Oc3ccc(C(F)(F)F)cn3)ccc2n1)N1CCN[C@@H](CO)C1
InChIKey: PLCWFJNNLUWLTR-OAHLLOKOSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: FRET-Based Membrane Potential Assay: Two days prior to the experiment, frozen HEK293 cells stably expressing recombinant human Nav1.7 were quickly thawed and plated at 25,000 cells/well in growth medium [DMEM (Invitrogen #11965) with 10% HI FBS (Invitrogen #10082), 2 mM glutamine, 100 units/mL penicillin, 0.1 mg/mL streptomycin (PSG, Sigma #G1146), and 500 ug/mL Geneticin (Invitrogen #10131)] in black-walled, clear-bottom 384-well poly-D-lysine-coated assay plates (Greiner Bio-One, Frickenhausen, Germany) and incubated in a humidified 5% CO2 incubator at 37° C. On the day of the assay, medium was removed by aspiration, and cells were washed with assay buffer [HBSS (Invitrogen, Carlsbad, Calif.) containing 20 mM HEPES (Invitrogen, Carlsbad, Calif.)]. After washing, 30 uL assay buffer containing the fluorescent voltage-sensor probe CC2-DMPE (Invitrogen, Carlsbad, Calif.) at 20 uM and 0.01% pluronic F-127 (Invitrogen, Carlsbad, Calif.) was added to the cells. Cells were incubated for 40 minutes at room temperature in the dark. Following the incubation, the cells were washed and 30 uL assay buffer containing 2.5 uM DiSBAC2(3) substrate (Invitrogen, Carlsbad, Calif.) and 0.5 mM VABSC-1 (Invitrogen, Carlsbad, Calif.) was added to the cells. The cells were incubated for 90 minutes at room temperature in the dark. Fluorescence readings were made using a FLIPR TETRA (Molecular Devices, Sunnyvale Calif.) equipped with voltage-sensor probe optics. At the start of each experiment the optimal (EC80) concentration of depolarizing agent (veratridine) was determined by testing a dilution curve of assay buffer containing veratridine (Sigma-Aldrich, St. Louis, Mo.) and 1 mg/mL scorpion venom (SVqq, from Leiurus quinquestriatus; Sigma-Aldrich, St. Louis, Mo.). Compounds were dissolved in dimethyl sulfoxide, and 8-point, 1:3 dilution concentration-response curves were prepared in duplicate in dimethyl sulfoxide, followed by preparation of 0.8 uL/well daughter plates of the dilutions. Test compounds in the daughter plate were diluted to (Ë¿3x) solutions in assay buffer immediately before assaying. Using the FLIPR TETRA, 20 uL of the (3x) compound solutions were first added to the cells, then 20 uL of depolarizing solution (3xEC80 veratridine+SVqq) were added 3 minutes later to activate the channel. Changes in fluorescence were measured at wavelengths of 440-480 nm and 565-625 nm over the course of the experimental run. Membrane depolarization was expressed as a ratio of the maximum F440-480 nm/F565-625 nm reading above average baseline F440-480 nm/F565-625 nm reading. IC50 values were calculated from curve fits of the ratio data using a four-parameter logistic Hill equation (Accelrys Assay Explorer 3.3 Client, Accelrys, San Diego, Calif.) with percent inhibition plotted against compound concentration.
Measurement: IC50 = 3120 nM (pChEMBL 5.510000228881836)
Descriptors: MW 432.4, ALogP 2.85, HBA 6, HBD 2, PSA 87.58, QED 0.66
Compound (SMILES): COc1ccc(CC(=O)O)cc1-c1ccc(F)c2c1CCN(C(=O)[C@@H]1C[C@H]1c1ccccc1)C2
InChIKey: BWYAGNFNCTXXMO-JTHBVZDNSA-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 = 414 nM (pChEMBL 6.380000114440918)
Descriptors: MW 459.52, ALogP 4.82, HBA 3, HBD 1, PSA 66.84, QED 0.57
Compound (SMILES): O=C1NCCc2[nH]c(-c3cccc4ccc(N[C@H]5CCCNC5)nc34)cc21
InChIKey: FXKAJMTVSSOIMV-AWEZNQCLSA-N
Target: Serine/threonine-protein kinase pim-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P11309
Assay: Homogeneous Time Resolved Fluorescence Assay (HTRF): The assay for the determination of Pim activity is based on the formation of phosphorylated biotinylated-BAD peptide at the Serine 112 residue (S112) and employs HTRF® (homogeneous time resolved fluorescence) technology to detect the product in a 96-well plate format. The phosphorylation of biotinylated-BAD (S112) peptide by full length recombinant Pim-1, Pim-2, or Pim-3 protein was detected with streptavidin:Allophycocyanin (APC) conjugate and a europium (Eu) labeled antibody directed against phosphorylated-BAD (S112). Excitation of Eu by a high energy laser light (337 nm) leads to a transfer of energy to the APC molecule, and results in an emission at 665 nm. The fluorescence is directly proportional to the amount of phosphorylated BAD peptide present in the reaction. Compounds were prepared in DMSO by conducting 3-fold serial dilutions to give a 10-point dosing curve having a high dose of 1 μM. A reference compound was included on each assay plate in order to validate that plate; on one plate of every assay run, two additional reference compounds were included. The final buffer conditions were as follows: 60 mM Hepes, pH 7.0, 0.05% BSA, 2 mM DTT. Incubations were carried out at RT (22° C.) for 2 h for Pim-1, 1 hour and 30 min for Pim-3, and 45 min for Pim-2. The reaction was stopped by the addition of 3 mM EDTA, and fluorescence was measured by an HTRF® Rubystar microplate reader.
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 361.45, ALogP 2.68, HBA 4, HBD 4, PSA 81.84, QED 0.58
Compound (SMILES): C[C@@]12[C@@H](CN(c3ccc(C#N)c(C(F)(F)F)c3)S1(=O)=O)[C@H]1C[C@H](O)[C@@H]2C1
InChIKey: MSTUMOQHZHYIEH-RLPSOPLKSA-N
Target: Androgen receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P10275
Assay: Antagonist activity at androgen receptor in human MDA-MB-453 cells assessed as inhibition of testosterone-induced transactivation
Measurement: IC50 = 106 nM (pChEMBL 6.96999979019165)
Descriptors: MW 386.39, ALogP 2.5, HBA 4, HBD 1, PSA 81.4, QED 0.8
Compound (SMILES): C[C@@H](c1ccccc1)[C@H](CN)N1CCN(c2ncnc(N)c2-c2ccc(F)cc2)CC1
InChIKey: AFTITNGEJQLJMI-UWJYYQICSA-N
Target: Ribosomal protein S6 kinase beta-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23443
Assay: Enzyme Assay: P70S6K inhibitor compounds were diluted and plated in 96 well plates. A reaction mixture including the following components was then added to the compound plate to initiate the enzyme reaction; P70S6K (3 nM, T412E mutant, Millipore) was mixed with 24 μM ATP in an assay buffer containing 100 mM Hepes (pH 7.5), 5 mM MgCl2, 1 mM DTT, 0.015% Brij and 1 μM of the substrate peptide FITC-AHA-AKRRRLSSLRA-OH (derived from the S6 ribosomal protein sequence, FITC=fluorescein isothiocyanate, AHA=6-aminohexanoic acid). The reaction was incubated for 90 min at 25° C., before the addition of 10 mM EDTA to stop the reaction.
Measurement: IC50 = 280 nM (pChEMBL 6.550000190734863)
Descriptors: MW 420.54, ALogP 3.12, HBA 6, HBD 2, PSA 84.3, QED 0.64
Compound (SMILES): Cc1cccc2cc(Cn3nc(I)c4c(N)ncnc43)n(-c3ccccc3F)c(=O)c12
InChIKey: WRUZUACCMTZECI-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P48736
Assay: Inhibition of recombinant human C-terminal His6-tagged p110gamma expressed in insect cells using phosphatidylinositol as substrate in presence of gamma-32P-ATP after 5 mins by thin layer chromatographic method
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 526.31, ALogP 3.81, HBA 7, HBD 1, PSA 91.62, QED 0.36
Compound (SMILES): Cc1nnc2ccc(C(=O)N3CCC[C@H]3c3ccc(Cl)cc3)cn12
InChIKey: UJIAQMCWYMQXCS-INIZCTEOSA-N
Target: CDK8/Cyclin C (PROTEIN COMPLEX), Homo sapiens
UniProt: P24863
Assay: Inhibition of Alexa647 tracer binding to full length recombinant human His-tagged CDK8/cyclin C expressed in Baculovirus expression system preincubated for 20 mins followed by tracer addition and incubated in dark for 60 mins by FRET-based Lanthascreen assay
Measurement: IC50 = 3990 nM (pChEMBL 5.400000095367432)
Descriptors: MW 340.81, ALogP 3.67, HBA 4, HBD 0, PSA 50.5, QED 0.71
Compound (SMILES): O=C(N[C@H]1CC[C@H](CCN2CCN(c3nsc4ccccc34)CC2)CC1)c1ccccc1
InChIKey: VZKXQURASTWIAW-AQYVVDRMSA-N
Target: 5-hydroxytryptamine receptor 1A (SINGLE PROTEIN), Homo sapiens
UniProt: P08908
Assay: Displacement of [3H]8-OH-DPAT from recombinant human 5-HT1A receptor expressed in HEK293 cell membranes after 60 mins by scintillation counting method
Measurement: Ki = 1.7 nM (pChEMBL 8.770000457763672)
Descriptors: MW 448.64, ALogP 4.8, HBA 5, HBD 1, PSA 48.47, QED 0.59
Compound (SMILES): O=C1[C@H]2CCC[C@H]2N1c1ccc(C#Cc2ccccc2)cn1
InChIKey: ZRRFNKLAOMQOSE-DLBZAZTESA-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 = 26 nM (pChEMBL 7.579999923706055)
Descriptors: MW 288.35, ALogP 3, HBA 2, HBD 0, PSA 33.2, QED 0.6
Compound (SMILES): COc1cc2c(Nc3ccc4occc4c3)ncnc2cc1OCCCCCCC(=O)NO
InChIKey: MHNMJCPZRQMSAC-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of N-terminal GST-His6-thrombin cleavage site-fused human VEGFR2 (Asp805 to Val1356 residues) expressed in baculovirus expression system using biotinylated gastrin precursor at Tyr87 as substrate preincubated for 5 mins followed by substrate addition measured after 30 mins by DELFIA method
Measurement: IC50 = 1000 nM (pChEMBL 6)
Descriptors: MW 450.5, ALogP 4.96, HBA 8, HBD 3, PSA 118.74, QED 0.17
Compound (SMILES): CC1=C(C(=O)Nc2ccccc2Cl)C(c2ccccc2Br)N=C(Nc2nc3ccccc3o2)N1
InChIKey: DSFMTARMUAMBBK-UHFFFAOYSA-N
Target: Galactokinase (SINGLE PROTEIN), Homo sapiens
UniProt: P51570
Assay: qHTS Assay: Assay details and protocol: The primary assay monitored ATP depletion using Promega's KinaseGlo technology, where ATP levels are measured through luminescence generated from firefly luciferase, a bioluminescent ATP-dependent enzymes. ATP was held at 35 μM, near its reported KM value, and the KM for galactose was determined under the 1536-well assay conditions to be 50-100M (FIG. 1A). As well, the IC50 for a commercially available CD45 inhibitor (N-(9,10-dioxo-9,10-dihydrophenanthren-2-yl)pivalamide) was confirmed (previously found to inhibit GALK (FIG. 1B)). This was used as the positive control for the assay. The assay used 5 nM GALK and a 1 hr incubation time, which gave sufficient signal:background and stability for the HTS.
Measurement: IC50 = 3260.9 nM (pChEMBL 5.489999771118164)
Descriptors: MW 536.82, ALogP 6.27, HBA 6, HBD 3, PSA 91.55, QED 0.29
Compound (SMILES): CCN1CCN(C(=O)c2cccc(Cc3n[nH]c(=O)c4c3CCCC4)c2)CC1
InChIKey: ZXFNUHYGGNERBC-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P09874
Assay: PARP Enzyme Inhibition Assay: PARP1 assay was conducted in PARP assay buffer containing 50 mM Tris pH 8.0, 1 mM DTT, 4 mM MgCl2. PARP reactions contained 1.5 uM [3H]-NAD+ (1.6 uCi/mmol), 200 nM biotinylated histone H1, 200 nM sIDNA, and 1 nM PARP enzyme. Auto reactions utilizing SPA bead-based detection were carried out in 100 uL volumes in white 96 well plates. Reactions were initiated by adding 50 ul of 2xNAD+ substrate mixture to 50 uL of 2x enzyme mixture containing PARP and DNA. These reactions were terminated by the addition of 150 uL of 1.5 mM benzamide (~1000-fold over its IC50). 170 uL of the stopped reaction mixtures were transferred to streptavidin Flash Plates, incubated for 1 hour, and counted using a TopCount microplate scintillation counter.
Measurement: Ki = 10 nM (pChEMBL 8)
Descriptors: MW 380.49, ALogP 2.02, HBA 4, HBD 1, PSA 69.3, QED 0.88
Compound (SMILES): Cl.NC(N)=N/N=C/c1ccc(-c2ccccc2)cc1
InChIKey: ZOWSXLOOOAEFET-LZMXEPDESA-N
Target: Heme oxygenase (SINGLE PROTEIN), Pseudomonas aeruginosa
UniProt: O69002
Assay: Binding affinity to Pseudomonas aeruginosa HemO by intrinsic fluorescence quenching method
Measurement: Kd = 6700 nM (pChEMBL 5.170000076293945)
Descriptors: MW 238.29, ALogP 1.96, HBA 2, HBD 2, PSA 76.76, QED 0.49
Compound (SMILES): Cn1cc(S(=O)(=O)NCc2cccc(C3(Cc4ccccc4)CNC(=O)C3)c2)cn1
InChIKey: GJNGATNTYBDWQM-UHFFFAOYSA-N
Target: Sodium- and chloride-dependent glycine transporter 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P48067
Assay: Radioligand Binding Assay: Radioligand binding to human GlyT1c transporter-expressing membranes was determined as described in Mezler et al., Molecular Pharmacology 74:1705-1715, 2008.
Measurement: Ki < 10 nM (pChEMBL 8)
Descriptors: MW 424.53, ALogP 1.9, HBA 5, HBD 2, PSA 93.09, QED 0.61
Compound (SMILES): COc1ccc([C@H](NC(=O)c2ccc3cnc(NC4CC(OC)C4)nc3c2)c2cnn(C)c2)cc1F
InChIKey: FTQPIABVIHXJIM-IMQCZEGASA-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 100× 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 μM S/T17 peptide substrate and 25 μM ATP (corresponding to the observed Km) for a total reaction volume of 25 μL. 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 = 13.8 nM (pChEMBL 7.860000133514404)
Descriptors: MW 490.54, ALogP 3.62, HBA 8, HBD 2, PSA 103.19, QED 0.39
Compound (SMILES): N#Cc1cccc(-c2ccc(/N=N/c3cc(S(=O)(=O)[O-])c4ccccc4c3N)cn2)c1.[Na+]
InChIKey: MRMAWOCOGQKVRL-JGUILPGDSA-M
Target: Transitional endoplasmic reticulum ATPase (SINGLE PROTEIN), Mus musculus
UniProt: Q01853
Assay: Inhibition Assay: Mouse VCP cDNA was added with a DNA sequence corresponding to histidine tag at the amino-terminal, subcloned into a baculovirus vector pVL1392 (BD Bioscience), and expressed in Sf-9 insect cells. The protein was purified with a nickel column (GE Healthcare). The concentration of the protein was adjusted to 0.25-0.5 ug/ml and the protein was stored in a solution containing 50 mM TrisCl pH 8.0, 5 mM EDTA, 10% glycerol, and 15 mM DTT. The ATPase activity was determined as follows. 500 ng of the purified VCP was mixed with 100 uM [gamma-32P]ATP (18.5 GBq/mmol) and the test substance in 20 uL of ATPase buffer (20 mM HEPES (pH7.4), 50 mM KCl, 5 mM MgCl2, 15 mM DTT), and incubated at 37° C. for 10 minutes.The enzyme reaction was stopped with addition of 200 uL of an ice-cold solution containing 7% TCA and 1 mM K2HPO4. 50ul of a solution containing 3.75% ammonium molybdate and 0.02M tungstate silicic/3 NH2SO4 was added, followed by 300 uL of n-butylacetic acid.
Measurement: IC50 = 584 nM (pChEMBL 6.230000019073486)
Descriptors: MW 429.46, ALogP 5.02, HBA 7, HBD 2, PSA 141.79, QED 0.26
Compound (SMILES): C[C@]12CCC(=O)C=C1C1CC1[C@@H]1[C@@H]2CC[C@]2(C)C(c3cccnc3)=CC[C@@H]12
InChIKey: FRSPDRIARDOSKB-ODCXVSDJSA-N
Target: Steroid 17-alpha-hydroxylase/17,20 lyase (SINGLE PROTEIN), Homo sapiens
UniProt: P05093
Assay: Inhibition of CYP17 in human testicular microsomes using 17alpha-hydroxypregnenolone as substrate assessed as decrease in DHEA levels at 1 nM after 90 mins by ELISA relative to control
Measurement: IC50 > 100 nM (pChEMBL 7)
Descriptors: MW 359.51, ALogP 5.46, HBA 2, HBD 0, PSA 29.96, QED 0.67
Compound (SMILES): COc1ccc(-c2ncn(CC(=O)N3CCCCC3)c2-c2ccc(OC)cc2)cc1
InChIKey: YFEMRANIMKQPFX-UHFFFAOYSA-N
Target: Prostaglandin G/H synthase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35354
Assay: Inhibition of human recombinant COX-2 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 = 14120 nM (pChEMBL 4.849999904632568)
Descriptors: MW 405.5, ALogP 4.25, HBA 5, HBD 0, PSA 56.59, QED 0.62
Compound (SMILES): CC(C)(O)CNC(=O)N1C(=O)N(c2cccc(C(F)(F)F)c2)C2=C(C(=O)CC2)[C@@H]1c1ccc(C#N)cc1
InChIKey: DOVSNSDMRJVTMR-QFIPXVFZSA-N
Target: Neutrophil elastase (SINGLE PROTEIN), Homo sapiens
UniProt: P08246
Assay: Neutrophil Elastase Assay: Materials: Human neutrophil elastase was purchased from Calbiochem (Cat. No.: 324681) and the elastase substrate MeOSuc-Ala-Ala-Pro-Val-AMC from Bachem (Cat. No.: I-1270). All other materials were of the highest grade commercially available. The following buffers were used: Compound buffer: 100 mM Tris, 500 mM NaCl, adjusted to pH 7.5; Assay buffer: 100 mM Tris, 500 mM NaCl, adjusted to pH 7.5, containing 0.01% BSA. Assay conditions: Test compounds were prediluted in DMSO and subsequently in compound buffer (5% DMSO final). 5 uL of these compound dilutions were mixed with 10 ul Neutrophil elastase (9 ng/ml in assay buffer) in a black 384 well OptiPlate (Perkin Elmer, Cat No.: 6007270) and incubated for 15 min at room temperature. Subsequently 10 uL substrate solution in assay buffer were added (250 uM final concentration) and the plates were incubated for 60 min at room temperature. After inactivation of the enzyme, fluorescence intensities were measured at 380 nm excitation and 460 nm emission wavelengths. Each plate contains wells with a high value control (DMSO+enzyme+substrate) and wells with a low value control (DMSO+inactivated enzyme+substrate). IC50 values were estimated using a sigmoidal concentration response curve with variable slope.
Measurement: IC50 = 40.4 nM (pChEMBL 7.389999866485596)
Descriptors: MW 512.49, ALogP 4.66, HBA 5, HBD 2, PSA 113.74, QED 0.62
Compound (SMILES): CN[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N1Cc2ccccc2[C@H]1C(=O)Nc1c(F)cccc1F
InChIKey: OHOQSMXCQQKXFR-LVWPNOBMSA-N
Target: E3 ubiquitin-protein ligase XIAP (SINGLE PROTEIN), Homo sapiens
UniProt: P98170
Assay: TR-FRET Assay: The peptide AVPIAQKSEK-(ε-biotin)-OH 1:2 TFA ("Peptide A") was identified as a substrate for the TR-FRET assay by screening the 6× Histidine-tagged BIR2 domain and BIR3 domain of XIAP against a set of 29 peptides synthesized based on sequences reported by Sweeny et al. (Biochemistry, 2006, 45, 14740 14748). The peptides were labeled with the fluorescent tags FITC or TAMRA and Kd values were determined by fluorescence polarization assay. The sequence AVPIAQKSEK was identified as optimal for using in an assay. The peptide sequence was derivatized with biotin to provide AVPIAQKSEK-(Îµ-biotin)-OH 1:2 TFA as the substrate for the TR-FRET assay. Ten nanomolar of 6× Histidine-tagged BIR2 domain, corresponding to amino acids 124-240 of XIAP, or BIR3 domain, corresponding to amino acids 241-356 of XIAP, was mixed with 20 nM of the peptide AVPIAQKSEK-(ε-biotin)-OH 1:2 TFA, in the presence of 50 mM Tris-Cl, 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 = 27 nM (pChEMBL 7.570000171661377)
Descriptors: MW 472.54, ALogP 3.13, HBA 4, HBD 3, PSA 90.54, QED 0.55
Compound (SMILES): COc1cc(OC)cc(N(CC#CC2CCN(C(=O)OC(C)(C)C)CC2)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: YOHMMSYTAZLPHD-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 5888.44 nM (pChEMBL 5.230000019073486)
Descriptors: MW 582.71, ALogP 5.84, HBA 9, HBD 0, PSA 94.84, QED 0.26