lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Nc1cncc(-c2cn3ccnc3c(Nc3ccc(N4CCN(C5COC5)CC4)c(OCCO)c3)n2)n1
InChIKey: RWEICHZKRDIUMB-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: High Throughput Syk Biochemical Assay: Syk activity was measured using KinEASE (Cisbio), a time-resolved fluorescence resonance energy transfer (TR-FRET) immunoassay. In this assay, Syk-catalyzes the phosphorylation of a XL665-labeled peptide substrate. Europium conjugated phospho-tyrosine specific antibody binds the resulting phosphorylated peptide. Formation of phosphorylated peptide is quantified by TR-FRET with Europium as the donor and XL665 the acceptor in a 2-step endpoint assay. In brief, test compounds serially diluted in DMSO were delivered into Corning white, low volume, non-binding 384 well plates using the Echo 550 acoustic liquid dispenser (Labcyte®). Syk enzyme and substrates were dispensed into assay plates using a Multi-Flo (Bio-Tek Instruments). The standard 5 μL reaction mixture contained 20 μM ATP, 1 μM biotinylated peptide, 0.015 nM of Syk in reaction buffer (50 mM Hepes, pH 7.0, 0.02% NaN3, 0.1% BSA, 0.1 mM Orthovanadate, 5 mM MgCl2, 1 mM DTT, 0.025% NP-40). After 30 minutes of incubati
Measurement: IC50 = 12.2 nM (pChEMBL 7.909999847412109)
Descriptors: MW 503.57, ALogP 1.4, HBA 12, HBD 3, PSA 139.19, QED 0.32
Compound (SMILES): CNC(=O)c1c(-c2ccc(F)cc2)oc2cc(N(CC(F)F)S(C)(=O)=O)c(-c3ccc4c(n3)-c3cc5c(C#N)cccc5n3CO4)cc12
InChIKey: JGIUEADGZIADLH-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1b (isolate Japanese) (HCV)
UniProt: P26662
Assay: ELISA-Based Assay: Measurement of inhibition by compounds was performed using the HCV replicon system. Several different replicons encoding different HCV genotypes or mutations were used. In addition, potency measurements were made using different formats of the replicon assay, including different ways of measurements and different plating formats. See Jan M. Vrolijk et al., A replicons-based bioassay for the measurement of interferons in patients with chronic hepatitis C, 110 J. VIROLOGICAL METHODS 201 (2003); Steven S. Carroll et al., Inhibition of Hepatitis C Virus RNA Replication by 2'-Modified Nucleoside Analogs, 278(14) J. BIOLOGICAL CHEMISTRY 11979 (2003). However, the underlying principles are common to all of these determinations, and are outlined below.Stable neomycin phosphotransferase encoding replicons-harboring cell lines were used, so all cell lines were maintained under G418 selection prior to the assay. Potency was determined using a cell ELISA assay with an antibody to the replicons encoded NS3/4a protease. See Caterina Trozzi et al., In Vitro Selection and Characterization of Hepatitis C Virus Serine Protease Variants Resistant to an Active-Site Peptide Inhibitor, 77(6) J. Virol. 3669 (2003).
Measurement: IC50 = 3.7 nM (pChEMBL 8.430000305175781)
Descriptors: MW 671.66, ALogP 6.53, HBA 8, HBD 1, PSA 130.46, QED 0.2
Compound (SMILES): CC1(C)CCC(CN2CCN(c3ccc(C(=O)NS(=O)(=O)c4ccc(NCC5COCCO5)c([N+](=O)[O-])c4)c(Oc4cccc5[nH]cnc45)c3)CC2)=C(c2ccc(Cl)cc2)C1
InChIKey: UZVXJUUTDBPGFF-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The inhibition constant (Ki) is the dissociation constant of an enzyme-inhibitor complex or a protein/small molecule complex, wherein the small molecule is inhibiting binding of one protein to another protein or peptide. Where the Ki for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-XL) is greater than the limits of detection of the assay used. Where the binding selectivity ratio for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the selectivity of a particular compound for Bcl-2 over Bcl-XL is at least as great as the number indicated. Where the Ki for a compound is represented as < (less than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-2) is lower than the limit of detection of the assay used. Inhibition constants were determined using Wang's equation (Wang Z-X).
Measurement: Ki = 32.06 nM (pChEMBL 7.489999771118164)
Descriptors: MW 870.43, ALogP 7.65, HBA 12, HBD 3, PSA 181.26, QED 0.08
Compound (SMILES): Cc1c(Br)cccc1Nc1ncc(Br)c(Nc2cc(C3CC3)[nH]n2)n1
InChIKey: HXQZIXCKQQGDSJ-UHFFFAOYSA-N
Target: Insulin-like growth factor 1 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08069
Assay: Inhibition of human IGF1R kinase domain (M954 to C1367 residues) tri-phosphorylated enzyme form expressed in baculovirus based expression system using poly-EY peptide incubated for 2 hrs by luciferase-luciferin coupled chemiluminescent assay
Measurement: IC50 = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 464.17, ALogP 5.4, HBA 5, HBD 3, PSA 78.52, QED 0.47
Compound (SMILES): Cc1cc(S(=O)(=O)N[C@H]2CCN([C@H](Cc3cc4ccnc(N)c4cc3OC(F)(F)F)C(=O)O)C2=O)cnc1N(C)C
InChIKey: PMQKOMRVCJPMLV-RBUKOAKNSA-N
Target: Coagulation factor X (SINGLE PROTEIN), Homo sapiens
UniProt: P00742
Assay: Inhibition Assay: The compounds according to the invention are tested in a range of concentrations (10pM to 10 µM final) in assay buffer (50 mM TRIS, 100 mM NaCl, 0.1 % BSA, pH 7,5) with 0.1% maximal final DMSO concentration and deposited in a proportion of 25 µl per well, on 25 µL enzyme (human coagulation factor Xa: Enzyme Research Laboratories HFXa, final concentration 0.003Ul/ml). The reactive were mixed, centrifuged and incubated 10 minutes at 37°C in a 96 well microtiter plate. The enzyme reaction was started with 50 µL substrate (S-2765, Biogenic ref 821413 in a final concentration of 62.5µM final). The time course of the reaction was monitored at 405 nm in a microtiter plate reader (Tecan M200) for 20 minutes at 37°C. The percentage of inhibition of enzymatic activity (expressed as maximum rate of cleavage of the substrate) is calculated with respect to the enzymatic activity in the absence of inhibitor.
Measurement: IC50 = 810 nM (pChEMBL 6.090000152587891)
Descriptors: MW 596.59, ALogP 2.06, HBA 9, HBD 3, PSA 168.05, QED 0.33
Compound (SMILES): CC[C@@H]1CN(c2nnc(Cc3ccccc3)c(C)c2C)CCN1c1cnc(C(C)(C)O)cn1
InChIKey: INLSIUHDZZBVKT-OAQYLSRUSA-N
Target: Protein smoothened (SINGLE PROTEIN), Mus musculus
UniProt: P56726
Assay: Inhibition of mouse recombinant Shh protein in mouse TMHh12 cells assessed as inhibition of 25 nM Smo agonist-induced hedgehog signalling response by Gli-luciferase reporter gene assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 446.6, ALogP 3.81, HBA 7, HBD 1, PSA 78.27, QED 0.62
Compound (SMILES): N=C(N)N[C@H]1CC[C@@H](C(=O)Nc2ccc(C[C@H](NC(=O)[C@@H]3CCCN3S(=O)(=O)c3ccccc3)C(=O)O)cc2)CC1
InChIKey: XFHCUQCLLGVBIK-ZXHYAQOVSA-N
Target: Integrin alpha-V/beta-1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P06756
Assay: Inhibition of integrin alphaVbeta1 (unknown origin)-mediated CHO cell adhesion to fibronectin preincubated for 15 to 30 mins followed by 60 min incubation for adhesion by crystal violet staining based cell adhesion assay
Measurement: IC50 = 1.995 nM (pChEMBL 8.699999809265137)
Descriptors: MW 584.7, ALogP 1.63, HBA 6, HBD 6, PSA 194.78, QED 0.18
Compound (SMILES): CC[C@H](C)[C@H](NC(=O)[C@H](CC1CCCCC1)NC(=O)c1cc(N)no1)C(=O)N1CCC2(C=Cc3ccccc32)CC1
InChIKey: KFVCQVXIFKORCI-QLADTWODSA-N
Target: Proteinase-activated receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P55085
Assay: Antagonist activity at PAR2 in human HT-29 cells assessed as inhibition of 2f-LIGRLO-NH2-stimulated intracellular calcium release preincubated for 30 mins followed by 2f-LIGRLO-NH2 stimulation measured for 60 sec by Fluo-3 AM dye based FLIPR assay
Measurement: IC50 = 1258.93 nM (pChEMBL 5.900000095367432)
Descriptors: MW 561.73, ALogP 4.44, HBA 6, HBD 3, PSA 130.56, QED 0.41
Compound (SMILES): CN1CC(NC(=O)N2C(=O)N(c3cccc(C(F)(F)F)c3)C3=C(C(=O)CC3)C2c2ccc(C#N)cc2)CC1=O
InChIKey: DPRBZPODWBUMNW-UHFFFAOYSA-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 < 1 nM (pChEMBL 9)
Descriptors: MW 537.5, ALogP 4.12, HBA 5, HBD 1, PSA 113.82, QED 0.63
Compound (SMILES): COCCOc1ccc2[nH]cc(C(=O)OCC34CCN(CC3)CC4)c2c1C
InChIKey: MSENZMOJTGKXAU-UHFFFAOYSA-N
Target: Neuronal acetylcholine receptor subunit alpha-7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q05941
Assay: Alpha-Bungarotoxin Binding Assay: Protocol 2: Studies were carried out using rat brain P2 membrane. All membrane was obtained from male Sprague-Dawley (SD) rats and collected on phosphate buffer. Protein concentration was determined using the bicinchoninic acid (BCA) protein assay reagent. In an α-Bungarotoxin binding assay, membrane was diluted with assay buffer (50 mM potassium phosphate, 1 mM pH 8.0 EDTA, 0.1 mM PMSF, 0.1% BSA). Test compounds were prepared in 100% DMSO solution. The starting concentration of the test compound was 3 μM, 8 points in 3 fold dilution. Assays were performed in a total volume of 200 μL containing [3H] α-Bungarotoxin (final concentration 0.4 nM), membrane suspension (80 μL containing 150 μg of membrane protein) and 20 μL of test compound. Non-specific binding (NSB) was determined with 1 μM α-Bungarotoxin. After 3 hours incubation (37° C.), the assay solution was transferred to a GF/B plate (Millipore). The assay was terminated by adding 100 μL ice-cold PBS dilution and then rapidly filtering over glass fiber filters presoaked in 0.5% polyethylenimine. The filters were washed 4 times with 200 μL/well of ice-cold PBS. They were then were transferred to the reading plate, and 250 μL of microscine 40 was added. The radioactivity trapped by the filters was determined by liquid scintillation counting MicroBeta.
Measurement: IC50 = 220.6 nM (pChEMBL 6.659999847412109)
Descriptors: MW 372.47, ALogP 3.14, HBA 5, HBD 1, PSA 63.79, QED 0.6
Compound (SMILES): Cc1ccc(C2=C(C(=O)Nc3ccccc3)C(=O)NC(c3ccc(OCCCC(F)(F)F)cc3)(C(F)(F)F)C2)cc1
InChIKey: WOTADNPUQOERCL-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: MGAT SPA Assay: MGAT2 enzyme was assayed using membranes isolated from Sf9 cells expressing the recombinant human MGAT2 cDNA with 2-monooleoylglycerol and [3H]-oleoyl-CoA as substrates as described by Seethala et al. [Anal. Biochem., 383(2):144-150 (Dec. 15, 2008)]. Briefly, the assays were conducted in 384-well plates in a total volume of 30 uL at 25° C. In each assay, 200 ng of recombinant human MGAT2 membrane was incubated with 10 uM of 2-monooleoylglycerol and 15 uM of [3H]-oleoyl-CoA in 100 mM potassium phosphate (pH 7.4) for 20 min with various concentrations of compounds delivered in DMSO. The assay was terminated by the addition of 20 ul of Stopping Solution (7.5 mg/ml Yttrium Oxide Polylysine beads, 3.3 mg/ml Fraction V BSA and 200 uM Mercuric chloride in 50 mM HEPES, pH 7.4). The signal was measured 1 h after quenching the reaction using LEADSEEKERSM for 5 minutes. To calculate the degree of inhibition, the zero level of enzyme activity (blank) was defined.
Measurement: IC50 = 53 nM (pChEMBL 7.28000020980835)
Descriptors: MW 576.54, ALogP 7.09, HBA 3, HBD 2, PSA 67.43, QED 0.17
Compound (SMILES): CCCOc1cc(N2CCC(N3CCCCC3)CC2)ccc1Nc1nccc(-c2c(-c3cccc(C(=O)Nc4c(F)cccc4F)c3)nc3ccccn23)n1
InChIKey: GJTVFKMEJTVXEL-UHFFFAOYSA-N
Target: Insulin receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P06213
Assay: Inhibition of human GST-rTEV-tagged insulin receptor (979 to 1382 residues) expressed in baculovirus infected insect Sf9 cells assessed as decrease in biotin-aminohexyl-AEEEEYMMMMAKKKK-NH2 phosphorylation after 1 hr by fluorometer
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 742.87, ALogP 8.98, HBA 9, HBD 2, PSA 99.92, QED 0.14
Compound (SMILES): Cn1ncc2ccc(OCC(=O)NCC(O)CN3CCc4ccccc4C3)cc21
InChIKey: JJSMEMABSCDVRO-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 5 (SINGLE PROTEIN), Homo sapiens
UniProt: O14744
Assay: Inhibition of PRMT5 in human Z138 cells assessed as decrease in di-methyl arginine levels after 96 hrs by Western blot method
Measurement: EC50 = 101 nM (pChEMBL 7)
Descriptors: MW 394.48, ALogP 1.49, HBA 6, HBD 2, PSA 79.62, QED 0.63
Compound (SMILES): O=S(=O)(Nc1nccs1)c1ccc2c(c1)OCCN2c1cccc2ccncc12
InChIKey: YIYIHOOPTATGMR-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: In Vitro PX Assay: HEK 293 cells stably transfected with human Nav1.7 were recorded in whole cell voltage clamp mode with the PatchXpress automated electrophysiology system (Molecular Devices, LLC, Sunnyvale, Calif.). Compound effects were measured on a partially inactivated state of the sodium channel. Cells were clamped to a holding potential yielding 20 to 50% inactivation. To elicit sodium current, channels were activated by pulsing to −10 mV for 20 msec. This voltage protocol was repeated at a rate of 0.1 Hz throughout the experiment. A single concentration of test compound was applied to cells for a duration of 3 minutes.
Measurement: IC50 = 1750 nM (pChEMBL 5.760000228881836)
Descriptors: MW 424.51, ALogP 4.02, HBA 7, HBD 1, PSA 84.42, QED 0.53
Compound (SMILES): CC(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(=O)N[C@H](C(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@H](C(=O)N[C@@H](CC(=O)O)C(=O)NCC(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@@H](Cc1ccccc1)C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](Cc1ccccn1)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(N)=O)C(C)C)C(C)C
InChIKey: KGZKWUODLMVMPO-YLACCPEKSA-N
Target: Tissue factor pathway inhibitor (SINGLE PROTEIN), Homo sapiens
UniProt: P10646
Assay: Binding affinity to TFPI (unknown origin) incubated for 1.5 hrs using tracer peptide JBT0131 by screening ELISA
Measurement: IC50 = 31 nM (pChEMBL 7.510000228881836)
Descriptors: MW 2407.76
Compound (SMILES): COc1ccnc(N2CCN(c3nccc4c(Cl)c([C@]5(C)CS(=O)(=O)N(C)C(=N)N5)sc34)CC2)c1
InChIKey: JTSNCCGBWXNHAI-QFIPXVFZSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: HTRF FRET Assay: Inhibitor compounds, prepared at 3× the desired final concentration in 1×BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO are pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1×BACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30° C. The assay is initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 μM for 4 μM for autoBACE-2) prepared in 1×BACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C. DMSO is present at 5% final concentration in the assay. Following laser excitation of sample wells at 320 nm, the fluorescence signal at 620 nm is collected for 400 ms following a 50 μs delay on a RUBYstar HTRF plate reader (BMG Labtechnologies).
Measurement: Ki = 1262 nM (pChEMBL 5.900000095367432)
Descriptors: MW 536.08, ALogP 2.69, HBA 9, HBD 2, PSA 114.75, QED 0.52
Compound (SMILES): O=C(O)c1ccc(CC[C@H]2C(=O)CC[C@@H]2/C=C/C(O)Cc2cccc(Cl)c2)cc1
InChIKey: YQAHEMJLXLEJPE-FSKZSRKQSA-N
Target: Prostaglandin E2 receptor EP2 subtype (SINGLE PROTEIN), Homo sapiens
UniProt: P43116
Assay: Radioligand Binding Assay: The cell homogenate was centrifuged at 19000 r.p.m. for 20 min at 4° C. using a Beckman Ti-60 rotor. The resultant pellet was resuspended in TME buffer to give a final 1 mg/ml protein concentration, as determined by Biorad assay. Radioligand binding competition assays vs. [3H-]17-phenyl PGF2a , (5 nM) were performed in a 100 μl volume for 60 min. Binding reactions were started by adding plasma membrane fraction. The reaction was terminated by the addition of 4 ml ice-cold TRIS-HCl buffer and rapid filtration through glass fiber GF/B filters using a Brandel cell harvester. The filters were washed 3 times with ice-cold buffer and oven dried for one hour. [3H-] PGE2 (specific activity 180 Ci mmol) was used as the radioligand for EP receptors. [3H] 17-phenyl PGF2a, was employed for FP receptor binding studies. Binding studies employing EP1, EP2, EP4 and FP receptors were performed in duplicate in at least three separate experiments. A 200 μl assay volume was used. Incubations w
Measurement: Ki = 2755 nM (pChEMBL 5.559999942779541)
Descriptors: MW 412.91, ALogP 4.73, HBA 3, HBD 2, PSA 74.6, QED 0.61
Compound (SMILES): CC(=O)c1cnc(N2CCN(c3nnc(Cc4ccccc4)c4c3CCC4)C[C@H]2C)nc1C(F)(F)F
InChIKey: UMYIUACAFHCDRE-MRXNPFEDSA-N
Target: Protein smoothened (SINGLE PROTEIN), Mus musculus
UniProt: P56726
Assay: Inhibition of mouse recombinant Shh protein in mouse TMHh12 cells assessed as inhibition of 25 nM Smo agonist-induced hedgehog signalling response by Gli-luciferase reporter gene assay
Measurement: IC50 = 100 nM (pChEMBL 7)
Descriptors: MW 496.54, ALogP 4.28, HBA 7, HBD 0, PSA 75.11, QED 0.49
Compound (SMILES): O=S(=O)(Nc1ncns1)c1ccc2c(c1)OCCN2c1ccc(C(F)(F)F)cc1-c1cn[nH]c1
InChIKey: ISRFCSFRFHSLEY-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: In Vitro PX Assay: HEK 293 cells stably transfected with human Nav1.7 were recorded in whole cell voltage clamp mode with the PatchXpress automated electrophysiology system (Molecular Devices, LLC, Sunnyvale, Calif.). Compound effects were measured on a partially inactivated state of the sodium channel. Cells were clamped to a holding potential yielding 20 to 50% inactivation. To elicit sodium current, channels were activated by pulsing to −10 mV for 20 msec. This voltage protocol was repeated at a rate of 0.1 Hz throughout the experiment. A single concentration of test compound was applied to cells for a duration of 3 minutes.
Measurement: IC50 = 57.6 nM (pChEMBL 7.239999771118164)
Descriptors: MW 508.51, ALogP 4.28, HBA 8, HBD 2, PSA 113.1, QED 0.41
Compound (SMILES): CNC(=O)CCc1ccc(Nc2nccc(Nc3cc(C4CC4)n[nH]3)n2)cc1
InChIKey: YYZUDSKOQRUJAH-UHFFFAOYSA-N
Target: Insulin-like growth factor 1 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08069
Assay: Inhibition of human IGF1R kinase domain (M954 to C1367 residues) tri-phosphorylated enzyme form expressed in baculovirus based expression system using poly-EY peptide incubated for 2 hrs by luciferase-luciferin coupled chemiluminescent assay
Measurement: IC50 = 500 nM (pChEMBL 6.300000190734863)
Descriptors: MW 377.45, ALogP 3.24, HBA 6, HBD 4, PSA 107.62, QED 0.48
Compound (SMILES): C[C@@]1(c2cc(N[C@H]3CCC[C@H]3O)ccc2F)COCC(N)=N1
InChIKey: RWCZTUDBYGQJTK-LZWOXQAQSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: Cellular Assay: The assay uses the principle of inhibition of human TMEM27 cleavage by endogenous cellular BACE2 in the Ins1e rat cell line and shedding from the cell surface into the culture medium, followed by detection in an ELISA assay. Inhibition of BACE2 prevents the cleavage and shedding in a dose-dependent mannerThe stable cell line "INS-TMEM27" represents an INS1e-derived cell line with inducible expression (using the TetOn system) of full-length hTMEM27 in a doxycycline-dependent manner. The cells are cultured throughout the experiment in RPMI1640 +Glutamax (Invitrogen) Penicillin/Streptomycin, 10% Fetal bovine serum, 100 mM pyruvate, 5 mM beta-mercatptoethanol, 100 micrograms/ml G418 and 100 microgram/ml hygromycin and are grown inadherent culture at 37° C. in a standard CO2 cell culture incubator.INS-TMEM27 cells are seeded in 96-well plates. After 2 days in culture, BACE2 inhibitor is added in a range of concentrations as required by the assay and after a further two hours, doxycycline i
Measurement: IC50 = 2270 nM (pChEMBL 5.639999866485596)
Descriptors: MW 307.37, ALogP 1.75, HBA 5, HBD 3, PSA 79.87, QED 0.79
Compound (SMILES): CCCCN(CCCC)C(=O)c1cc(C)n(-c2ccc(N[S+]([O-])c3sc(Cl)cc3Br)cc2C(=O)N2Cc3ccccc3C[C@H]2CO)n1
InChIKey: BCFHUKMNHHSXDA-POQAZEJGSA-N
Target: Apoptosis regulator Bcl-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P10415
Assay: Surface plasmon resonance (SPR): Methods for Determining IC50s.The present method includes utility of a Surface plasmon resonance (SPR)- based biosensor (Biacore GE Healthcare, Uppsala, Sweden) to characterize BCL-2 inhibitors.Biacore utilizes the phenomenon of surface plasmon resonance (SPR) to detect and measure binding interactions. In a typical Biacore experiment, one of the interacting molecules (ligand) is immobilized on a flexible dextran matrix while the interacting partner (analyte) is allowed to flow across that surface. A binding interaction results in an increase in mass on the sensor surface and a corresponding direct change in the refractive index of the medium in the vicinity of the sensor surface. Changes in refractive index or signal are recorded in resonance units (R.U.) Signal changes due to association and dissociation of complexes are monitored in a non-invasive manner, continuously and in real-time, the results of which are reported in the form of a sensorgram.
Measurement: IC50 = 465.17 nM (pChEMBL 6.329999923706055)
Descriptors: MW 761.21, ALogP 7.39, HBA 8, HBD 2, PSA 113.76, QED 0.14
Compound (SMILES): NC(=O)/C=C/c1c[nH]c2ncnc(Oc3ccccc3)c12
InChIKey: PCGJEVAVUOYIQB-VOTSOKGWSA-N
Target: TAK1/TAB1 (PROTEIN COMPLEX), Homo sapiens
UniProt: Q15750
Assay: Inhibition of kinase tracer-236 binding to full length recombinant human His-tagged TAK1-TAB1 expressed in baculovirus by FRET assay
Measurement: IC50 = 170 nM (pChEMBL 6.769999980926514)
Descriptors: MW 280.29, ALogP 2.25, HBA 4, HBD 2, PSA 93.89, QED 0.72
Compound (SMILES): CN1Cc2c(Cl)cc(Cl)cc2[C@H](c2cccc(S(=O)(=O)NCCOCCOCCOCCn3cc(CN(Cc4cn(CCOCCOCCOCCNS(=O)(=O)c5cccc([C@@H]6CN(C)Cc7c(Cl)cc(Cl)cc76)c5)nn4)Cc4cnnn4CCOCCOCCOCCNS(=O)(=O)c4cccc([C@@H]5CN(C)Cc6c(Cl)cc(Cl)cc65)c4)nn3)c2)C1
InChIKey: KIZXFRAJMQPROD-UCBZISERSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P48764
Assay: Inhibition of human NHE3-mediated sodium-dependant hydrogen ion antiport in Opossum kidney cells preincubated in NH4Cl-HEPES buffer containing BCECF-AM dye followed by washout with ammonium free HEPES and subsequent addition of compound in presence of NHE1 antagonist EIPA by fluorescence assay
Measurement: IC50 = 501.19 nM (pChEMBL 6.300000190734863)
Descriptors: MW 1848.72
Compound (SMILES): CC(=O)N1CCN(C(=O)N2CC[C@](c3ccc(C(OCc4c(F)cccc4C#N)(C(F)(F)F)C(F)(F)F)cc3)(S(=O)(=O)c3ccc(F)cc3)C2)CC1
InChIKey: QJQIXZIPDHUXKU-HKBQPEDESA-N
Target: Nuclear receptor ROR-gamma (SINGLE PROTEIN), Homo sapiens
UniProt: P51449
Assay: Inverse agonist activity at RORgammat in human Jurkat cells by Gal4 reporter assay
Measurement: EC50 = 17 nM (pChEMBL 7.769999980926514)
Descriptors: MW 758.69, ALogP 6.03, HBA 6, HBD 0, PSA 111.02, QED 0.22
Compound (SMILES): O=C1CN(c2ccccc2Cl)CCN1CC(O)CN1CCc2ccccc2C1
InChIKey: SCOHWISZLDYORY-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 5 (SINGLE PROTEIN), Homo sapiens
UniProt: O14744
Assay: Inhibition of PRMT5 in human Z138 cells assessed as decrease in di-methyl arginine levels after 96 hrs by Western blot method
Measurement: EC50 = 101 nM (pChEMBL 7)
Descriptors: MW 399.92, ALogP 2.41, HBA 4, HBD 1, PSA 47.02, QED 0.84
Compound (SMILES): O=C(Nc1ccncn1)N1CCN(Cc2ccc3c(c2)OC(F)(F)O3)CC1
InChIKey: IKKLWYSRDQOURR-UHFFFAOYSA-N
Target: Fatty-acid amide hydrolase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O00519
Assay: FAAH Assay: T84 frozen cell pellets or transfected SK-N-MC cells (contents of 1x15 cm culture dishes) were homogenized in 50 mL of FAAH assay buffer (125 mM Tris, 1 mM EDTA, 0.2% Glycerol, 0.02% Triton X-100, 0.4 mM Hepes, pH 9). The assay mixture consisted of 50 uL of the cell homogenate, 10 uL of the test compound, and 40 uL of anandamide [1-3H-ethanolamine] (3H-AEA, Perkin-Elmer, 10.3 Ci/mmol), which was added last, for a final tracer concentration of 80 nM. The reaction mixture was incubated at rt for 1 h. During the incubation, 96-well Multiscreen filter plates (catalog number MAFCNOB50; Millipore, Bedford, Mass., USA) were loaded with 25 uL of activated charcoal (Multiscreen column loader, catalog number MACL09625, Millipore) and washed once with 100 uL of MeOH. Also during the incubation, 96-well DYNEX MicroLite plates (catalog number NL510410) were loaded with 100 uL of MicroScint40 (catalog number 6013641, Packard Bioscience, Meriden, Conn., USA).
Measurement: IC50 = 60000 nM (pChEMBL 4.21999979019165)
Descriptors: MW 377.35, ALogP 2.15, HBA 6, HBD 1, PSA 79.82, QED 0.88
Compound (SMILES): COC(=O)[C@H](C)Nc1cc(C(=O)O)nc(-c2ccc(Oc3ccc(F)cc3)cc2)n1
InChIKey: WCRBNOREWVLNLP-LBPRGKRZSA-N
Target: N-formyl peptide receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P25090
Assay: FLIPR Assay: Assay Buffer: The assay buffer was formulated by removing 120 mL from a 1 L bottle of fresh, sterile dH2O (Mediatech, Herndon, Va.) and adding 100 mL of 10×HBSS that does not contain Ca++ or Mg++ (Gibco, Invitrogen, Grand Island, N.Y.) followed by 20 mL of 1.0 M Hepes, pH 7.3 (Fisher Scientific, BP299-100). The final buffer consisted of 20 mM Hepes, pH 7.3, 1.261 mM CaCl2, 0.493 mM MgCl2, 0.407 mM Mg(SO)4, 5.33 mM KCl, 0.441 mM KH2PO4, 137 mM NaCl, 0.336 mM Na2HPO4 and 0.556 mM D-glucose (Hanks et al., Proc. Soc. Exp. Biol. Med. 71:196 (1949)), and the simple formulation was typically the basic buffer throughout the assay (i.e., all wash and addition steps).
Measurement: IC50 = 970 nM (pChEMBL 6.010000228881836)
Descriptors: MW 411.39, ALogP 3.75, HBA 7, HBD 2, PSA 110.64, QED 0.57
Compound (SMILES): CC(C)(C)C(CO)NC(=O)c1cnc(C2CC2)c(OCC2CC2)n1
InChIKey: KWNJQZSSKRFKMZ-UHFFFAOYSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1×HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37° C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30° C. for 30 min. Compounds were added to a final assay volume of 100 μl and incubated for 30 min at 30° C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 μl lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN3) and 50 μl detection solutions (20 μM mAb Alexa700-cAMP 1:1, and 48 μM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 333.43, ALogP 2.28, HBA 5, HBD 2, PSA 84.34, QED 0.8
Compound (SMILES): COCCN1CCC(Nc2ccc(S(=O)(=O)NC(=O)c3ccc(N4CCN(CC5=C(c6ccc(Cl)cc6)CC(C)(C)CC5)CC4)cc3Oc3cccc4[nH]ccc34)cc2[N+](=O)[O-])CC1
InChIKey: YDBMCECCEUUCHH-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The inhibition constant (Ki) is the dissociation constant of an enzyme-inhibitor complex or a protein/small molecule complex, wherein the small molecule is inhibiting binding of one protein to another protein or peptide. Where the Ki for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-XL) is greater than the limits of detection of the assay used. Where the binding selectivity ratio for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the selectivity of a particular compound for Bcl-2 over Bcl-XL is at least as great as the number indicated. Where the Ki for a compound is represented as < (less than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-2) is lower than the limit of detection of the assay used. Inhibition constants were determined using Wang's equation (Wang Z-X).
Measurement: Ki = 32.03 nM (pChEMBL 7.489999771118164)
Descriptors: MW 910.54, ALogP 8.95, HBA 11, HBD 3, PSA 162.38, QED 0.07
Compound (SMILES): Cn1cncc1-c1ccnc2c1cnn2-c1ccc(F)cc1
InChIKey: JFEMPGINTWTZJM-UHFFFAOYSA-N
Target: Cytochrome P450 1A2 (SINGLE PROTEIN), Homo sapiens
UniProt: P05177
Assay: Inhibition of CYP1A2 (unknown origin)
Measurement: IC50 = 58 nM (pChEMBL 7.239999771118164)
Descriptors: MW 293.31, ALogP 2.96, HBA 5, HBD 0, PSA 48.53, QED 0.57
Compound (SMILES): NC(=O)c1cc(-c2ccc(F)cc2)c2ccc(CCc3ccccn3)cc2n1
InChIKey: UEUKQZXLBISYPI-UHFFFAOYSA-N
Target: Metabotropic glutamate receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q14416
Assay: Binding affinity to human mGluR2 expressed in cell membrane incubated for 1 hr in presence of tritium labeled [3H]-7-((2,5-dioxopyrrolidin-1-yl)methyl)-4-(4-fluorophenyl)quinoline-2-carboxamide by competitive radioligand binding assay
Measurement: Ki = 15 nM (pChEMBL 7.820000171661377)
Descriptors: MW 371.42, ALogP 4.32, HBA 3, HBD 1, PSA 68.87, QED 0.57
Compound (SMILES): CC[C@@H](CCCC(C)(C)O)N1C(=O)[C@@](C)(CC(=O)O)C[C@H](c2cccc(Cl)c2)C1c1ccc(Cl)cc1
InChIKey: VDEGZYFADJNHRX-WLSCKKSTSA-N
Target: E3 ubiquitin-protein ligase Mdm2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00987
Assay: Homogenous Time-Resolved Fluorescence Assay (HTRF2 Assay): The standard assay conditions for the in vitro HTRF assay consisted of a 50 ul total reaction volume in black 384-well Costar polypropylene plates in 1×PBS buffer pH 7.4, 1 mM DTT, 0.1% BSA, 2.5 nM GST-hMDM2 (aa 1-188), 5 nM biotinylated-p53 (aa 1-83), 1.8 nM SA-XLent (Cisbio; Bedford, Mass.), 0.6 nM anti-GST cryptate monoclonal antibody (Cisbio; Bedford, Mass.) and 200 mM KF. Amino acid residues 1-188 of human MDM2 were expressed as an amino-terminal glutathione-S-transferase (GST) fusion protein (GST-hMDM2) in Escherichia coli. Residues 1-83 of human p53 were expressed as an amino-terminal AviTag™-TrxA-6×His fusion protein (biotinylated p53) in E. coli. Each protein was purified from cell paste by affinity chromatography.Specifically, 10 uL of GST-hMDM2 was incubated with 10 ul of diluted compound (various concentrations, serially diluted) in 10% DMSO for 20 minutes at room temperature. 20 uL of biotinylated-p53 was added to the GST-hMDM2+compound mixture, and then incubated at room temperature for 60 min. 10 uL of detection buffer consisting of SA-XLent, anti-GST cryptate antibody and KF was added to GST-hMDM2, biotinylated-p53 and compound reaction and left at room temperature to reach equilibrium for >4 hrs. The final concentration of DMSO in the reaction was 2%. Time-resolved fluorescence readings were measured on a microplate multilabel reader. Percentage of inhibition was calculated relative to nutlin-3. All assay conditions remained the same as described above, with the exception of the following changes in reagent concentrations: 0.2 nM GST-hMDM2 (1-188), 0.5 nM biotinylated-p53 (1-83), 0.18 nM SA-XLent, and 100 mM KF.
Measurement: IC50 = 4 nM (pChEMBL 8.399999618530273)
Descriptors: MW 534.52, ALogP 7.25, HBA 3, HBD 2, PSA 77.84, QED 0.34
Compound (SMILES): Cc1cc2ccc1[C@@H](C)COC(=O)Nc1ccc(C3(c4nnn[nH]4)CC3)c(c1)CN(C)C(=O)[C@@H]2Nc1ccc2c(N)nccc2c1
InChIKey: YSEANFNTDGIIGS-AFJIDDCJSA-N
Target: Coagulation factor VII (SINGLE PROTEIN), Homo sapiens
UniProt: P08709
Assay: In Vitro Assay: Tissue kallikrein-1 (37° C) activity was determined in reactions containing 0.05 nM enzyme and 90 μM substrate (H-D-Val-Leu-Arg-AFC) in buffer (0.1M sodium phosphate pH 7.4, 0.2 M NaCl, 0.5% PEG 8000, and 1% DMSO). Assays were performed using 96-well microtiter plates (COSTAR 3600, CORNING, NY, USA) and a thermostatic temperature controlled plate reader (SPECTRAMAX® Gemini, Molecular Devices, Sunnyvale, Calif., USA). Fluorescence was monitored using 400 nm excitation and 505 nm emission wavelengths.
Measurement: Ki = 230 nM (pChEMBL 6.639999866485596)
Descriptors: MW 617.71, ALogP 5.2, HBA 9, HBD 4, PSA 164.04, QED 0.21
Compound (SMILES): COc1nc2ccc(C(C)(O)c3cnc(C)n3C)cc2c(Cl)c1Cc1ccc(C(F)(F)F)cc1
InChIKey: ZSGWUOZOUGDWCX-UHFFFAOYSA-N
Target: Nuclear receptor ROR-gamma (SINGLE PROTEIN), Homo sapiens
UniProt: P51449
Assay: ThermoFluor® Assay: ThermoFluor® experiments were carried out using instruments owned by Janssen Research and Discovery, L.L.C. through its acquisition of 3-Dimensional Pharmaceuticals, Inc. 1,8-ANS (Invitrogen) was used as a fluorescent dye. Protein and compound solutions are dispensed into black 384-well polypropylene PCR microplates (Abgene) and overlayed with silicone oil (1 μL, Fluka, type DC 200) to prevent evaporation. Bar-coded assay plates are robotically loaded onto a thermostatically controlled PCR-type thermal block and then heated at a typical ramp-rate of 1° C./min for all experiments. Fluorescence was measured by continuous illumination with UV light (Hamamatsu LC6) supplied via fiber optic and filtered through a band-pass filter (380-400 nm; >6 OD cutoff). Fluorescence emission of the entire 384-well plate was detected by measuring light intensity using a CCD camera (Sensys, Roper Scientific) filtered to detect 500±25 nm, resulting in simultaneous and independent readings of all 384 wells.Â  Reference wells contained RORγt without compounds, and the assay conditions were as follows: 0.065 mg/mL RORγt, 60 μM 1,8-ANS, 100 mM Hepes, pH 7.0, 10 mM NaCl, 2.5 mM GSH, 0.002% Tween-20.
Measurement: Kd = 470 nM (pChEMBL 6.329999923706055)
Descriptors: MW 489.93, ALogP 5.8, HBA 5, HBD 1, PSA 60.17, QED 0.38
Compound (SMILES): O=c1cc(/C=C/c2ccccc2)c2cc(C(c3ccc(Cl)cc3)c3ccc(Cl)cc3)ccc2[nH]1
InChIKey: VGJKOWCTRTVIKA-VOTSOKGWSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 482.41, ALogP 8.19, HBA 1, HBD 1, PSA 32.86, QED 0.25
Compound (SMILES): O=C1COc2ccc(CN3CCC(c4nnn5cnc6[nH]ccc6c45)CC3)cc2N1
InChIKey: MLEBNVKKLBPPHJ-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 200 nM (pChEMBL 6.699999809265137)
Descriptors: MW 403.45, ALogP 2.32, HBA 7, HBD 2, PSA 100.44, QED 0.54
Compound (SMILES): Cc1cc([N+](=O)[O-])cnc1N1CCc2nc(C(C)C)nc(Nc3ccc(C(F)(F)F)cc3)c2CC1
InChIKey: RSAZQLBLGVJNJC-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: O35433
Assay: Functional Assay: The assay was performed as described below, using HEK293 cells transfected with rat TRPV1 (rTRPV1/HEK293). These cells had a geneticin selection marker and were grown in Dulbecco's Modified Eagle Medium (DMEM, Gibco BRL) supplemented with 10% fetal bovine serum, penicillin/streptomycin (50 units/mL), and 500 μg/mL geneticin in 5% CO2 at 37° C. Cells were passaged frequently, every 3-5 days, to avoid overgrowth, depletion of essential medium components, or acidic medium exposure. Cells were passaged using a brief wash in 0.05% trypsin with 1 mM EDTA, followed by dissociation in divalent-free phosphate-buffered saline (Hyclone #SH30028.02). Dissociated cells were seeded onto poly-D-lysine coated black-walled 96-well plates (Biocoat; Becton Dickinson #354640) at about 40,000 cells per well and grown for approximately 1 day in culture medium to near confluency. The assay buffer was composed of 130 mM NaCl, 2 mM KCl, 2 mM MgCl2, 10 mM HEPES, 5 mM glucose, and either 2 mM or 20 μM CaCl2. On the day of the experiment, the culture medium was replaced with 2 mM calcium assay buffer using an automated plate washer (ELx405; Biotek, VT). The cells were incubated in 100 μL/well Fluo-3/AM (2 μM; TEFLabs #0116) with Pluronic F127 (100 μg/mL; Sigma #P2443) for 1 h at rt in the dark. After loading the cells, the dye solution was replaced with 50 μL/well of 20 μM calcium assay buffer using the ELx405 plate washer. Test compounds (50 μL/well) were added to the plate and incubated for 30 min. Intracellular Ca2+ levels were subsequently assayed using a Fluorometric Imaging Plate Reader (FLIPR™ instrument, Molecular Devices, CA) to simultaneously monitor Fluo-3 fluorescence in all wells (λexcitation=488 nm, λemission=540 nm) during challenge with agonist (capsaicin). The IC50 values were determined. Cells were challenged with 150 nM capsaicin and the fluorescence counts were captured following agonist addition at a sampling rate of 0.33 Hz. The contents of the wells were mixed 3 times (40 μL mix volume) immediately after the additions were made. Concentration dependence of block was determined by exposing each well of cells in duplicate rows of a 96 well plate to a serial dilution of test compound. The concentration series usually started at 10 Î¼M with a three-fold serial decrement in concentration. The magnitude of the capsaicin response was determined by measuring the change in fluo3 fluorescence before and 100 seconds after the addition of the agonist.
Measurement: IC50 = 350 nM (pChEMBL 6.460000038146973)
Descriptors: MW 486.5, ALogP 5.58, HBA 7, HBD 1, PSA 97.08, QED 0.37
Compound (SMILES): O=C(CCCCCCn1cc(-c2ccc3ncnc(Nc4ccc(OCc5nccs5)c(Cl)c4)c3c2)nn1)NO
InChIKey: GQWVZXYZPPWMNU-UHFFFAOYSA-N
Target: Histone deacetylase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13547
Assay: Inhibition of HDAC1 (unknown origin) using acetylated peptide substrate preincubated for 15 mins followed by substrate addition measured after 60 mins by fluorescence analysis
Measurement: IC50 = 5.6 nM (pChEMBL 8.25)
Descriptors: MW 579.09, ALogP 5.78, HBA 11, HBD 3, PSA 139.97, QED 0.09
Compound (SMILES): COc1cc(-c2ccc(Cl)c(C)c2)c(F)cc1-n1c(=O)c(OC)cc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: OPPSMXCUGDWMAJ-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Nav1.7 In Vitro PX Assay: HEK 293 cells stably transfected with human Nav1.7 were recorded in whole cell voltage clamp mode with the PatchXpress automated electrophysiology system (Molecular Devices, LLC, Sunnyvale, Calif.). Compound effects were measured on a partially inactivated state of the sodium channel. Cells were clamped to a holding potential yielding 20 to 50% inactivation. To elicit sodium current, channels were activated by pulsing to -10 mV for 20 msec. This voltage protocol was repeated at a rate of 0.1 Hz throughout the experiment. A single concentration of test compound was applied to cells for a duration of 3 minutes.
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 570, ALogP 5.56, HBA 8, HBD 1, PSA 112.66, QED 0.27
Compound (SMILES): CC(C)(C)OC(=O)N1CCC(CC#N)(n2cc(C(N)=O)c(Nc3cc(Cl)nc(Cl)c3)n2)CC1
InChIKey: YGTIBEJMKHOJJP-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: HTRF Biochemical Assay: The reactions employed a common peptide substrate, LCB-EQEDEPEGDYFEWLW-NH2 (in-house). The basic assay protocol is as follows: First, 250 nL of diluted compounds in DMSO were dispensed into the wells of a dry 384-well Black plate (Greiner #781076) using a Labcyte Echo 555 acoustic dispenser. Subsequent reagent additions employed an Agilent Bravo. Next, 18 uL of 1.11x enzyme and 1.11x substrate in 1x assay buffer (Invitrogen kinase buffer # PV3189, 2 mM DTT, 0.05% BSA) were added to the wells and shaken and then preincubated for 30 minutes at room temperature to allow compound binding to equilibrate. After equilibration, 2 uL of 10xATP in 1x assay buffer was added to initiate the kinase reaction and the plates were shaken and then incubated at room temperature for 120 minutes. At the end of the incubation, 20 uL of 2x stop buffer (streptavidin-Dylight 650 (Thermo #84547B/100 mL), Eu-tagged pY20 antibody (Perkin Elmer #AD0067), EDTA, HEPES, and Triton) was added to quench the reaction.
Measurement: IC50 = 33 nM (pChEMBL 7.480000019073486)
Descriptors: MW 494.38, ALogP 4.07, HBA 8, HBD 2, PSA 139.16, QED 0.6
Compound (SMILES): Cc1c(C(=O)Nc2cccc(C(F)(F)F)c2)cc(C(C)(C)C)n1C1CCN(C2CCNCC2)CC1
InChIKey: NPLKALZIQZJQGG-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9HBA0
Assay: Inhibition Assay: Test Example 1: (1) The TRPV4 inhibitory activity was measured using CHO-K1 cells stably expressing human TRPV4 (hTRPV4/CHO cells). (2) In the day before the experimental day, frozen cell were thawed and washed with the culture medium (MEM-α, 10% FBS, 2 mmol/L GlutaMax, 50 unit Penicillin, 50 μg/mL Streptomycin). Then cells were suspended in the culture medium. (3) The 384-well plates which hTRPV4/CHO cells were seeded at densities of 4000 cells/well in the culture medium and cultured in a CO2 incubator in the presence of 5% CO2 at 37° C. for overnight was used as assay plates. (4) The assay plate was washed with assay buffer (Hanks, 20 mmol/L HEPES, 2.5 mmol/L probenecid, pH7.4), and the buffer were remained at 20 μL/well. (5) 10 μl of dye loading buffer (9 μmol/L Fluo 4-AM, 0.09% Pluronic F-127/assay buffer) was added to each well, then the assay plate was incubated in a CO2 incubator in the presence of 5% CO2 at 37° C. for one hour. (final 3 μmol/L Fluo4-AM). (6) The assay plate was washed with the assay buffer, and the buffers were remained at 20 μL/well. Then the assay plate was incubated for 10 min at room temperature. (7) Diluted compound solution was dispensed to each well in the compound plate, and mixed with built-in Pipette and Mixer of the Fluorescence analysis system FLIPR TETRA (Molecular devices). (8) After incubation for 5 min, 20 μL of 4α-PDD solution were applied to each well of assay plate and mixed with the FLIPR TETRA. (final 1 Î¼mol/L 4α-PDD). (9) The fluorescent intensity was measured with FLIPR TETRA system for 10 min from the point of time addition the compound solution, at Ex 470-495 nm, Em 515-575 nm wavelength.
Measurement: IC50 = 6300 nM (pChEMBL 5.199999809265137)
Descriptors: MW 490.61, ALogP 5.75, HBA 4, HBD 2, PSA 49.3, QED 0.57
Compound (SMILES): CC1(C)OC(=O)N(C2CCC(c3cnc4[nH]ccc4c3)CC2)[C@H]1c1ccccc1
InChIKey: BCWOZUOLDWFOGA-LFNLJPBCSA-N
Target: Poly [ADP-ribose] polymerase tankyrase-1 (SINGLE PROTEIN), Homo sapiens
UniProt: O95271
Assay: Tankyrase Biochemical Assays: The tankyrase 1 biochemical activity of the compounds was assayed in the following assay buffer (50 mM MOPS pH7.5, 100 mM NaCl, 2.5 mM MgCl2, 0.01% Tween-20, 0.05% BSA, and 1 mM DTT) as follows: 0.25 nM of 6×HIS-tankyrase 1 (1091-1325) was incubated in the presence of compound (DMSO 1.85% final) in a Perkin Elmer 384 well Proxiplate Plus™ (cat. no. 6008289) with 400 nM of NAD for 60 minutes at room temperature. The assay was then stopped with the above assay buffer containing a 0.6 M inhibitor and the following detection components: 0.05 g/mL monoclonal anti-PAR antibody (Trevigen cat. no. 4335-MC-01K-AC) prebound for 60 minutes with 0.63 μg/mL protein G AlphaLisa® acceptor bead (Perkin Elmer cat. no. AL102M) and 5 μg/mL AlphaLisa® nickel chelate donor bead (Perkin Elmer cat. no. AS101M). The assay was incubated for 16 hours at room temperature in the dark and read on a Perkin Elmer Envision® multi label reader using the default program set with laser excitation at 680 nM and emission at 615 nM. The tankyrase 2 biochemical assay was performed using the procedure set forth above with respect to tankyrase 1 except that 4 nM 6×HIS-tankyrase 2 (946-1162) and 250 nM NAD were used.
Measurement: IC50 = 218 nM (pChEMBL 6.659999847412109)
Descriptors: MW 389.5, ALogP 5.56, HBA 3, HBD 1, PSA 58.22, QED 0.64
Compound (SMILES): CCOc1ccc(-c2nc3ccccn3c2-c2ccnc(Nc3ccc(N4CCC(N5CCC(CCS(C)(=O)=O)CC5)CC4)cc3OC)n2)cc1C(=O)Nc1c(F)cccc1F
InChIKey: JJMALFMQOBLFLD-UHFFFAOYSA-N
Target: Insulin receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P06213
Assay: Inhibition of human GST-rTEV-tagged insulin receptor (979 to 1382 residues) expressed in baculovirus infected insect Sf9 cells assessed as decrease in biotin-aminohexyl-AEEEEYMMMMAKKKK-NH2 phosphorylation after 1 hr by fluorometer
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 865.02, ALogP 8.26, HBA 12, HBD 2, PSA 143.29, QED 0.11