lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): N#Cc1ccc(C2C3=C(CCC3=O)N(c3cccc(C(F)(F)F)c3)C(=O)N2C(=O)NC2CC[S+]([O-])CC2)cc1
InChIKey: GSLXONFDAAJEEW-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 556.57, ALogP 4.8, HBA 5, HBD 1, PSA 116.57, QED 0.55
Compound (SMILES): CCOc1ccc(C)cc1C(=O)N[C@H]1CCC[C@@H]1Nc1cnc(C(F)(F)F)cn1
InChIKey: XJJSVHDKBXHBLI-GJZGRUSLSA-N
Target: Orexin/Hypocretin receptor type 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O43613
Assay: Fluorescent Imaging Plate Reader (FLIPR) Assay: Orexin antagonist activity was determined by measuring changes in intracellular calcium levels using a Ca2+ sensitive fluorescent dye. The changes in fluorescent signal were monitored by Fluorescent Imaging Plate Reader (FLIPR®) technology available from Molecular Devices, LLC, U.S.A. Orexin mediated increases in intracellular Ca2+ concentration were readily detected upon activation with orexin-A. Twenty-four hours prior to the assay, RBL-2H3 cells stably expressing either human orexin receptor 1 or human orexin receptor 2 were seeded in cell culture medium in black, clear-bottom 384-well plates (commercially available from Corning Inc., U.S.A.) and grown overnight at 37 °C., 5% CO2. On the day of the assay, cell culture media was removed and cells were loaded with Calcium 5 Dye (commercially sold by Molecular Devices, LLC, U.S.A.) for 1 hour at 37 °C., 5% CO2.
Measurement: IC50 = 279 nM (pChEMBL 6.550000190734863)
Descriptors: MW 408.42, ALogP 3.97, HBA 5, HBD 2, PSA 76.14, QED 0.76
Compound (SMILES): NC(=O)[C@@H]1N(C(=O)c2ccc(N3CC(F)(F)C3)c(OCC3CC3)n2)CCC12CC2
InChIKey: UOCYXIBLTUGJNB-HNNXBMFYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1×HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37 °C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30 °C. for 30 min. Compounds were added to a final assay volume of 100 μl and incubated for 30 min at 30 °C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 μl lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN3) and 50 μl detection solutions (20 μM mAb Alexa700-cAMP 1:1, and 48 μM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated as follows: FRET=T730-Alexa730-P(T645-B645) with P=Ru730-B730/Ru645-B645, where T730 is the test well measured at 730 nM, T645 is the test well measured at 645 nm, B730 and B645 are the buffer controls at 730 nm and 645 nm, respectively. cAMP content is determined from the function of a standard curve spanning from 10 μM to 0.13 nM cAMP.
Measurement: EC50 = 818 nM (pChEMBL 6.090000152587891)
Descriptors: MW 406.43, ALogP 1.81, HBA 5, HBD 1, PSA 88.76, QED 0.78
Compound (SMILES): CC(C)[C@H]1C[C@@H](c2ccccc2)CCN1C(=O)[C@H]1CNC[C@@]12CCCc1ccccc12
InChIKey: ZUIUWQIQNSLVIX-HDEUIXPWSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Electrochemiluminescence (ECL) Assay: A homogeneous end point electrochemiluminescence (ECL) assay is performed using a biotinylated BACE substrate. The Km of the substrate is approximated at 50 uM. A typical reaction contains approximately 0.6 nM enzyme, 0.25 uM of the substrate, and buffer (50 mM Pipes, pH 6.5, 0.1 mg/ml BSA, 0.2% CHAPS, 15 mM EDTA and 1 mM deferoxamine) in a total reaction volume of 100 ul. The reaction proceeds for 1-2 hrs and is then stopped by the addition of 150 uL of a quench cocktail solution (25 ul M Tris-HCl, pH 8.0, 50 ul INC buffer (2% BSA, 0.2% Tween-20 and 0.05% sodium azide diluted in Phosphate buffered saline (PBS) plus 75 uL PBS), containing Streptavidin coated magnetic beads and ruthenylated antibody which specifically recognizes the C-terminal residue of the product. The samples are subjected to M-384 (Igen Inc., Gaithersburg, MD) analysis. Under these conditions, less than 10% of substrate is processed by BACE 1.
Measurement: IC50 = 22000 nM (pChEMBL 4.659999847412109)
Descriptors: MW 416.61, ALogP 4.91, HBA 2, HBD 1, PSA 32.34, QED 0.77
Compound (SMILES): N[C@H]1CC[C@H](Nc2nc(NC3CC3)n3ncc(/C=C4\NC(=O)NC4=O)c3n2)CC1
InChIKey: NHXZVVIDPXHKJF-GHCBGBTPSA-N
Target: Casein kinase II subunit beta (SINGLE PROTEIN), Homo sapiens
UniProt: P67870
Assay: Inhibition Assay: 5 uM ATP: Test compounds dissolved and diluted in DMSO (2 μl) were added to a reaction mixture comprising 10 μl of 5× Reaction Buffer (40 mM MOPS pH 7.0, 5 mM EDTA), 10 μl of recombinant human PIM2 solution (4 ng PIM-2 dissolved in dilution buffer (20 mM MOPS pH 7.0; EDTA 1 mM; 5% Glycerol; 0.01% Brij 35; 0.1%; 0.1% 2-mercaptoethanol; 1 mg/ml BSA)) and 8 ul of water. Reactions were initiated by the addition of 10 ul of ATP Solution (49% (15 mM MgCl2; 5 uM ATP) 1% ([γ-33P]ATP: Stock 1 mCi/100 μl; 3000 Ci/mmol (Perkin Elmer)) and 10 ul of substrate peptide solution (RSRSSYPAGT (SEQ ID NO.: 6), dissolved in water at a concentration of 1 mM), Reactions were maintained for 10 min at 30° C. The reactions were quenched with 100 ul of 0.75% Phosphoric acid, then transferred to and filtered through a Phosphocellulose filter plate (Millipore, MSPH-N6B-50). After washing each well 4 times with 0.75% Phosphoric acid, scintillation fluid (20 uL) was added to each well and
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 397.44, ALogP 0.56, HBA 9, HBD 5, PSA 151.36, QED 0.36
Compound (SMILES): O=C(Nc1cc(OC2CCOC2)cc(C(F)(F)F)c1)Nc1ccc(Oc2ccnc(NC(=O)C3CC3)c2)c(F)c1
InChIKey: YTZXXSBOADXMTO-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition Assay: [Experimental method]: Use Lance@Ultra Ulight™-TK assay kit from PerKinElmer to evaluate the inhibitory activity of compounds at KDR kinase. [Instrument]: PerKinElmer's ENVISION plate reader. [Materials]: Optiplate-384 well plate (PerKinElmer), kinase buffer (50 mM Hepes pH7.5, 0.25 mM EGTA, 2 mM DTT, 0.01% Tween 20, 10 mM Mg2+, 0.5 mM Mn2+), KDR kinase (790-1356 AA, Crown Bioscience), KDR kinase substrate (PerKinElmer catalogue # TRF0127-M), Lance@Eu-W1024-anti-phosphotyrosine (PT66) (PerKinElmer, catalogue #AD0068), ATP (Invitrogen), DMSO (Sigma, catalogue #34869), purified water (Millipore, type: Milli-Q). [Study conditions]: Mix KDR (final concentration: 20 nM) and compound (final DMSO: 0.5%), pre-incubate for 20 min at 30° C.; then add ATP (final concentration: 90 μM) and the substrate (final concentration 50 μM). React for 2 hrs at 30° C. After the reaction, add antibody and reaction for 60 min at 30° C. Read the plate (615 nm, 665 nm), calculate the ratio of the value at 665 nm vs 615 nm and analyze the data.
Measurement: IC50 = 14.2 nM (pChEMBL 7.849999904632568)
Descriptors: MW 560.5, ALogP 6.19, HBA 6, HBD 3, PSA 110.81, QED 0.28
Compound (SMILES): CC(=O)N1CCc2nc(N3CCC(Oc4ccc(F)cc4F)CC3)c(NCC(F)F)nc2C1
InChIKey: RXQZHPDNUHUQAQ-UHFFFAOYSA-N
Target: G-protein coupled receptor 6 (SINGLE PROTEIN), Homo sapiens
UniProt: P46095
Assay: GPR6 In Vitro Assay (A.2): This cell based assay measures the ability of compounds to inhibit the constitutive cAMP activity of GPR6 receptor expressed in CHO-K1 cells. CHO cells were stably expressed with GPR6 receptor, whose expression is controlled by a tetracycline inducable element. The cells were cultured in medium containing F12K, 10% FBS, 1% PennStrep, 200 ug/mL Hygromycin. GPR6 receptor expression was induced for 20 hrs with 2 ug/ml doxycycline (sigma D9891) in growth media. After addition of doxycycline cells were plated at a density of 450-750 cells per well in 96-well half-volume black tissue culture plates (Costar) and placed in an incubator (37°, 5% CO2) for 20 hours prior to cAMP assays. Culture media was removed from cells and they were washed with 50 uL/well of Ringer's Buffer (MgCl2 0.047 mg/mL, NaH2PO4 0.18 mg/mL, Na2HPO4 0.1 mg/mL, KCl 0.34 mg/mL, NaHCO3 1.26 mg/mL, D-glucose 1.8 mg/mL, NaCl 7 mg/mL; pH=7.4). Compounds suspended in DMSO were diluted in Ringer's Buffer.
Measurement: IC50 = 0.0497 nM (pChEMBL 10.300000190734863)
Descriptors: MW 467.47, ALogP 3.38, HBA 6, HBD 1, PSA 70.59, QED 0.66
Compound (SMILES): COc1cc(C)c2[nH]ccc2c1C(C)(N)c1nc2cc(C#N)cnc2[nH]1
InChIKey: WSWGHGKCTBFCRQ-UHFFFAOYSA-N
Target: Complement factor B (SINGLE PROTEIN), Homo sapiens
UniProt: P00751
Assay: Biological Assay: Recombinant human factor B (expressed in drosophila cells and purified using standard methods) labeled with biotin (10 nM), europium-labeled streptavidin (5 nM) and (+) or (−)-2-((1E,3E,5E)-5-(1-(6-((2-(3-(4-((R)-3-amino-3-phenylpropanoyl)-1-(4-amino-6,7-dimethoxyquinazolin-2-yl)piperazin-2-yl)phenoxy)ethyl)amino)-6-oxohexyl)-3,3-dimethyl-5-sulfoindolin-2-ylidene)penta-1,3-dien-1-yl)-1-ethyl-3,3-dimethyl-5-sulfo-3H-indol-1-ium (Biological Example 2.6, 240 nM activity against factor B when tested using the assay of Biological Example 1) (75 nM) were incubated with test compound at various concentrations up to 2 hours at room temperature in 20 mM Tris/HCl, pH 7.4, 0.005% (v/v) Tween20.The time-gated decrease in fluorescence intensity related to the competition between labeled and unlabeled factor B ligands was recorded at both 620 nm and 665 nm, 70 μs after excitation at 337 nm using a microplate spectrofluorimeter.
Measurement: IC50 = 40 nM (pChEMBL 7.400000095367432)
Descriptors: MW 346.39, ALogP 2.85, HBA 5, HBD 3, PSA 116.4, QED 0.53
Compound (SMILES): COc1ccc(C2CC(c3ccc(F)cc3)=NN2c2ccc(C(=O)O)cc2)cc1
InChIKey: PFQJGOZMKDZHJQ-UHFFFAOYSA-N
Target: cGMP-specific 3',5'-cyclic phosphodiesterase (SINGLE PROTEIN), Homo sapiens
UniProt: O76074
Assay: Fluorescence Polarization Assay: PDE activity was measured using the IMAP fluorescence polarization assay (Molecular Devices) in which binding of hydrolyzed cyclic nucleotide substrate to immobilized metal coordination complexes increases fluorescence polarization (FP). Tetramethylrhodamine (TAMRA)-cGMP and fluorescein-cAMP were used as substrates, each at final concentration of 50 nmol/L. The PDE assay was done according to the manufacturer's specifications using either whole cell lysates or recombinant enzymes. FP was measured at excitation, emission wavelengths of either 530,590 nm for TAMRA-cGMP or 485,530 nm for fluorescein-cAMP using a Synergy4 (Biotek) microplate reader.
Measurement: IC50 = 3100 nM (pChEMBL 5.510000228881836)
Descriptors: MW 390.41, ALogP 4.89, HBA 4, HBD 1, PSA 62.13, QED 0.67
Compound (SMILES): CC(C)Oc1ccc(-c2nc(-c3cccc4c3CC[C@H]4N[C@H](C)c3ccccc3)no2)cc1C#N
InChIKey: JQMYJKHEYDAWDP-NIYFSFCBSA-N
Target: Sphingosine 1-phosphate receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21453
Assay: Agonist activity at human S1P1 expressed in CRE-bla CHOK1 cells assessed as inhibition of forskolin-induced cAMP formation preincubated for 4 hrs followed by FRET-based beta-lactamase fluorescent substrate addition measured after 2 hrs
Measurement: EC50 > 5 nM (pChEMBL 8.300999641418457)
Descriptors: MW 464.57, ALogP 6.4, HBA 6, HBD 1, PSA 83.97, QED 0.34
Compound (SMILES): CCN1CCN(c2ccc(Nc3ncc4c(C)c(C(C)=O)c(=O)n(C5CCCC5)c4n3)nc2)CC1
InChIKey: LICNBVSSOUPXNB-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00534
Assay: Inhibition of CDK6 (unknown origin) using histone H1 as substrate after 10 mins in presence of [gamma32P]ATP
Measurement: IC50 = 34 nM (pChEMBL 7.46999979019165)
Descriptors: MW 475.6, ALogP 3.7, HBA 9, HBD 1, PSA 96.25, QED 0.54
Compound (SMILES): Cc1ccccc1COc1ccc(S(=O)(=O)N2CCN(C(=O)CN)[C@@H](C)[C@@H]2C(=O)NO)cc1
InChIKey: LELBRQIPJQZJNJ-HRAATJIYSA-N
Target: Interstitial collagenase (SINGLE PROTEIN), Homo sapiens
UniProt: P03956
Assay: Inhibition of trypsin-activated recombinant human collagenase 1 preincubated with protein followed by addition of DNP-Pro-Cha-Gly-Cys(Me)-His-Ala-Lys(NMA)-NH2 as substrate measured after 20 mins by fluorescence analysis
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 476.56, ALogP 0.63, HBA 7, HBD 3, PSA 142.27, QED 0.39
Compound (SMILES): COc1ccc2nc(NC(=O)Nc3ccccc3N3CC4(CCN(CC(C)(C)C)CC4)c4c(F)ccc(O)c43)sc2c1
InChIKey: WPCZNZUAGNYTER-UHFFFAOYSA-N
Target: P2Y purinoceptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P47900
Assay: Scintillation Proximity Assay (SPA): A SPA membrane binding assay was used to identify inhibitors of [33P] 2MeS-ADP binding to cloned human P2Y1 receptors (The P2Y1 receptor membranes were provided by Biology and the cloning of the receptor and P2Y1 receptor membrane preparation is same as described by Biology). Binding reactions were performed in 384-well OptiPlates (PerkinElmer Life Sciences, Cat #6007299) in a volume of 50 uL containing 15 fmol of P2Y1 receptor (1.7 ug of total protein), 0.3 nM [33P] 2MeS-ADP (PerkinElmer; 2,000 Ci/mmol), various concentrations of the test compound (usually between 10 uM and 160 pM) in Buffer B containing 1% DMSO in assay buffer (15 mM, HEPES, 145 mM potassium chloride, 5 mM sodium Chloride, 5 mM EDTA, 0.1 mM MgCl2, pH 7.4) and 100 ug of SPA bead (WGA polystyrene Image beads, #RPNQ 0260V, Amersham). Reactions were allowed to proceed to completion at room temperature for 1 hour followed by centrifugation of the plate for 5 min. About 40 uL of the aqueous soluttion was aspirated. Plates were sealed and the [33P] 2MeS-ADP bound to the P2Y1 receptor membranes that were bound to the SPA bead were determined in a Gen 4 LEADSEEKERSM (Amersham) Image Reader. Dose-response curves (IC50) were fit by non-linear regression (Toolset an in house data processing program) and binding constants (Ki) calculated using the Cheng-Prusoff relationship (Ki=IC50/(1+L/Kd) in which a Kd for 2MeS-ADP to the P2Y1 receptor was determined to be 14 nM.
Measurement: Ki = 7.9 nM (pChEMBL 8.100000381469727)
Descriptors: MW 589.74, ALogP 7.33, HBA 7, HBD 3, PSA 89.96, QED 0.23
Compound (SMILES): COc1cc(-c2nc(-c3cc(C)nc(C)c3)c3n2CCOCC3)ccc1OC(F)F
InChIKey: OCKTURZLTOIAQU-UHFFFAOYSA-N
Target: Metabotropic glutamate receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q14416
Assay: [35S]GTPgammaS Binding Assay: The frozen cell membrane fraction prepared as described above was thawed before use, and the resultant was diluted with a buffer for a binding assay (final concentrations: 20 mM HEPES, 100 mM NaCl, 1 mM MgCl2, 3 uM GDP, 300 ug/mL saponin, 0.1% BSA). The compound of each example was added to a cell membrane fraction containing 1.8 to 3 ug/assay of membrane protein on a plate, followed by incubation at room temperature for 30 minutes. Thereafter, with glutamic acid (in a final concentration of 10 uM) added thereto, incubation was performed at room temperature for 15 minutes, and thereafter, [35S]GTPγS (in a final concentration of 0.8 kBq) and 588 ug WGA-SPA beads were added thereto, followed by incubation at room temperature for 1 hour. After the incubation, the plate was subjected to the centrifugal separation at 2,500 rpm and room temperature, and then, membrane cell binding radioactivity was measured with a top count. A [35S]GTPγS binding amount obtained in the presence of glutamic acid was defined as specific binding. On the basis of ratios of inhibiting the specific binding at various concentrations of the compounds of the respective examples, inhibition curves were obtained. Concentrations of the compounds of the respective examples at which the specific [35S]GTPγS binding amount was suppressed by 50% (IC50 values) were calculated on the basis of the inhibition curves.
Measurement: IC50 = 12.1 nM (pChEMBL 7.920000076293945)
Descriptors: MW 415.44, ALogP 4.41, HBA 6, HBD 0, PSA 58.4, QED 0.62
Compound (SMILES): Fc1ccc(-c2cnn(Cc3cc[nH]n3)c2)cc1OC(F)F
InChIKey: LHMLJRCNGQZNHL-UHFFFAOYSA-N
Target: Glutamate receptor ionotropic, NMDA 2B (SINGLE PROTEIN), Homo sapiens
UniProt: Q13224
Assay: Antagonist activity at human NR2B expressed in HEK293 cells co-expressing NRI assessed as inhibition of glutamate/glycine-induced Ca2+ flux measured after 90 secs
Measurement: IC50 = 370 nM (pChEMBL 6.429999828338623)
Descriptors: MW 308.26, ALogP 3.06, HBA 4, HBD 1, PSA 55.73, QED 0.79
Compound (SMILES): O=C(Nc1cccc(NC(=O)c2ccc(-c3cc(Cl)ccc3Cl)o2)c1)c1ccccc1
InChIKey: MKYNOIOLIDIFOA-UHFFFAOYSA-N
Target: Sentrin-specific protease 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P0U3
Assay: Inhibition of SENP1 catalytic domain (419 to 644 residues) (unknown origin) expressed in Escherichia coli BL21 (DE3) assessed as reduction in deSUMOylation preincubated for 10 mins followed by addition of RanGAP-SUMO as substrate measured after 25 mins by coomassie staining-based SDS-PAGE analysis
Measurement: IC50 = 17600 nM (pChEMBL 4.75)
Descriptors: MW 451.31, ALogP 6.76, HBA 3, HBD 2, PSA 71.34, QED 0.35
Compound (SMILES): O=C(Nc1cccc(CN2CCC(C(=O)NC3CCCC3)CC2)c1)c1ccc(F)cc1
InChIKey: VYIMDXFKDRCESX-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 0.9 nM (pChEMBL 9.050000190734863)
Descriptors: MW 423.53, ALogP 4.35, HBA 3, HBD 2, PSA 61.44, QED 0.73
Compound (SMILES): CCc1csc([C@H](Cc2ccc(NS(=O)(=O)O)cc2)NC(=O)Cc2ccc(F)c(F)c2)n1
InChIKey: AGKCTKMOEMXUNB-IBGZPJMESA-N
Target: Receptor-type tyrosine-protein phosphatase beta (SINGLE PROTEIN), Homo sapiens
UniProt: P23467
Assay: Inhibition of recombinant HPTPbeta (unknown origin) preincubated for 10 mins followed by substrate addition measured after 15 mins by DiFMUP probe-based fluorescence assay
Measurement: IC50 = 75 nM (pChEMBL 7.119999885559082)
Descriptors: MW 481.55, ALogP 3.84, HBA 5, HBD 3, PSA 108.39, QED 0.4
Compound (SMILES): Cc1ccc(C(=O)Nc2cccc(CN3CCC(C(=O)NC4CCCCC4)CC3)c2)c(Cl)n1
InChIKey: VNVMADASNDWPSB-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 40 nM (pChEMBL 7.400000095367432)
Descriptors: MW 469.03, ALogP 4.96, HBA 4, HBD 2, PSA 74.33, QED 0.59
Compound (SMILES): C[C@H]1COCCN1c1ncnc2[nH]cc(-c3cccc(C#N)c3)c12
InChIKey: XRIIKVLOTOSHOP-LBPRGKRZSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: Lantha Screen Technology Assay: LRRK2 kinase activity was measured using Lantha Screen technology from Invitrogen. GST-tagged truncated LRRK2 from Invitrogen (Cat # PV4874) was incubated with a fluorescein-labeled peptide substrate based upon ezrin/radixin/moesin (ERM), also known as LRRKtide (Invitrogen cat #PR8976A), in the presence of a dose response of compound. Upon completion, the assay was stopped and detected with a terbium labeled anti-phospho-ERM antibody (Invitrogen, cat #PR8975A). The assay was carried out under the following protocol: 3 uL of a working solution of substrate (233 nM LRRKtide, 117 uM ATP) prepared in assay buffer (50 mM HEEPES, pH 7.5, 3 mM MgCl2, with 2 mM DTT and 0.01% Brij35 added fresh) was added to a low volume Greiner 384-well plate. The compound dose response was prepared by diluting compound to a top concentration of 3.16 mM in 100% DMSO and serial diluted by half-log in DMSO 11 times. Aliquots (3.5 uL) of the 100% DMSO dose response were mixed with 46.5 uL water.
Measurement: IC50 = 44 nM (pChEMBL 7.360000133514404)
Descriptors: MW 319.37, ALogP 2.72, HBA 5, HBD 1, PSA 77.83, QED 0.79
Compound (SMILES): C#CC(C)N(Cc1cccc(-c2nccs2)c1)C(=O)Nc1ccc(OC(F)(F)F)cc1
InChIKey: LVSLJRGGEVZHSH-UHFFFAOYSA-N
Target: High affinity nerve growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P04629
Assay: Kinase Activity Assay: TrkA kinase activity was measured as the ability of the enzyme to phosphorylate a fluorescently labeled peptide substrate. Buffer salts, reagents, and fluorescently labeled peptides were purchased from Carna Biosciences and were of the highest quality available (QSS Assist TRKA_MSA Kit). Enzyme was purchased from Cell Signaling, and was used without further purification. 384-well round bottom assay plates were prepared by the addition of 200 nl of a DMSO solution of compound at various concentrations to final inhibitor concentrations ranging from 100 μM to 0.2 μM. Next, assay buffer (10 μl) containing substrate and ATP were added, followed by addition of 10 μl of enzyme in assay buffer. Final assays concentrations were: [E]=0.37 nM, [S]=1 μM, [ATP]=2 mM. The reactions were incubated at room temperature for 3 hours resulting in approximately 15% substrate phosphorylation and were terminated by the addition of 5 μl of stop buffer.
Measurement: EC50 = 48 nM (pChEMBL 7.320000171661377)
Descriptors: MW 445.47, ALogP 5.76, HBA 4, HBD 1, PSA 54.46, QED 0.49
Compound (SMILES): CC(C)Nc1cc(-n2ncc3cc(C#N)cnc32)ncc1C(=O)NCCc1ccncc1
InChIKey: KGNYBYULAFTJGU-UHFFFAOYSA-N
Target: Interleukin-1 receptor-associated kinase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NWZ3
Assay: Inhibition Assay: The assays were performed in U-bottom 384-well plates. The final assay volume was 30 L prepared from 15 L additions of enzyme and substrates (fluoresceinated peptide and ATP) and test compounds in assay buffer (20 mM HEPES pH 7.2, 10 mM MgCl2, 0.015% Brij 35 and 4 mM DTT). The reaction was initiated by the combination of IRAK4 with substrates and test compounds. The reaction mixture was incubated at room temperature for 60 min. and terminated by adding 45 μL of 35 mM EDTA to each sample. The reaction mixture was analyzed on the Caliper LABCHIP® 3000 (Caliper, Hopkinton, Mass.) by electrophoretic separation of the fluorescent substrate and phosphorylated product. Inhibition data were calculated by comparison to no enzyme control reactions for 100% inhibition and vehicle-only reactions for 0% inhibition. The final concentrations of reagents in the assays are ATP, 500 μM; FL-IPTSPITTTYFFFKKK peptide 1.5 μM; IRAK4, 0.6 nM; and DMSO, 1.6%.
Measurement: IC50 = 2.5 nM (pChEMBL 8.600000381469727)
Descriptors: MW 426.48, ALogP 2.87, HBA 8, HBD 2, PSA 121.41, QED 0.47
Compound (SMILES): CCC(=O)N1C[C@H](C)N(c2nc3cc(-c4nc(=O)o[nH]4)nc(-c4cncc(Cl)c4)c3n2C[C@H]2CC[C@H](C)CC2)[C@@H](C)C1
InChIKey: YEJOMGIETZPANP-MUGJNUQGSA-N
Target: E3 ubiquitin-protein ligase Mdm2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00987
Assay: FRET Assay: Methods: An HDM2 FRET assay was developed to assess the compounds' inhibitory activity towards binding of p53 protein. A truncated version of HDM2 with residues 17 to 125 (containing p53 binding surface, Science (1994) 265, 346-355), with N-terminal His and Thioredoxin tag was generated in pET32a expression vector and expressed in E. coli strain BL21(DE3)Rosetta. Protein was purified using Ni-affinity chromatography, followed by size exclusion chromatography using Superdex 75 26/60 column. To assess inhibition of p53 binding to HDM2, a FITC labeled 8-mer peptide (SEQ ID NO:1: Ac-Phe-Arg-Dpr-Ac6c-(6-Br)Trp-Glu-Glu-Leu-NH2; Anal Biochem. 2004 Aug. 1; 331(1):138-46) with strong affinity towards the p53 binding pocket of HDM2 was used. The HDM2 assay buffer contained 1x Phosphate Buffered Saline (Invitrogen, Cat#14190), 0.01% BSA (Jackson ImmunoResearch, Cat#001-000-162), 0.01% Tween-20.
Measurement: IC50 = 3 nM (pChEMBL 8.520000457763672)
Descriptors: MW 593.13, ALogP 5.15, HBA 9, HBD 1, PSA 126.04, QED 0.33
Compound (SMILES): Clc1ccc(CNC[C@H]2CC[C@H](c3nnn4cnc5[nH]ccc5c34)CC2)cc1
InChIKey: WVYXBGBSMGFRJF-KOMQPUFPSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
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 = 2.9 nM (pChEMBL 8.539999961853027)
Descriptors: MW 394.91, ALogP 4.32, HBA 5, HBD 2, PSA 70.9, QED 0.53
Compound (SMILES): Cc1nn(C)c(C)c1N(CC(F)F)S(=O)(=O)c1c(Cl)cc(CCCCC2CCNCC2)cc1Cl
InChIKey: NCFHKHWPNGZJEO-UHFFFAOYSA-N
Target: Glycylpeptide N-tetradecanoyltransferase (SINGLE PROTEIN), Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
UniProt: Q388H8
Assay: Enzyme Inhibition Assay: Measurements were performed using a modification of the scintillation proximity assay platform described previously by Georgopapadakou, N. H. et al. (22nd International Congress on Chemotherapy, 2001, Abstract P16.001). Compounds were solubilised in DMSO at a top concentration of 10 mM and serially diluted in half log steps to achieve a range of final assay concentrations of 100 uM to 1 nM. Compound at each concentration (100-fold final) was added to white 384 well plates in a volume of 0.5 ml. Human, A. fumigatus, T. brucei or L. major N-myristoyl transferase enzyme, dissolved to a working concentration of 10 nM in assay buffer (30 mM Tris/HCl pH 7.4, 0.5 mM EGTA, 0.5 mM EDTA, 1.25 mM DTT, 0.1% Triton X-100), was then added to columns 1 to 11 and 13 to 23 of the plates in a volume of 20 ml. To columns 12 and 24, 20 ml assay buffer was added to provide a no enzyme control. Following a 5 minute incubation at room temperature the substrates (GCGGSKVKPQPPQAK(Biotin)-Amide and myristoyl coenzyme A), dissolved in assay buffer, were added to all wells in a volume of 20 ml to start the reaction. The final concentrations of peptide and 3H-myristoyl coenzyme A were 0.5 mM and 125 nM respectively and the specific activity of the radiolabel was 8 Ci/mmol. Plates were then incubated at room temperature for up to 50 minutes (dependant upon the period of linearity for the different enzyme species) before SPA beads, suspended to 1 mg/ml in a stop solution (200 mM Phosphoric Acid/NaOH pH 4, 750 mM MgCl2), were added in a volume of 40 ml.
Measurement: IC50 = 3 nM (pChEMBL 8.520000457763672)
Descriptors: MW 537.5, ALogP 5.52, HBA 5, HBD 1, PSA 67.23, QED 0.41
Compound (SMILES): COc1ccc2cc(-c3n[nH]c4ccc(C(N)=O)cc34)ccc2c1
InChIKey: BLWTVYBQDIFPOQ-UHFFFAOYSA-N
Target: Mitogen-activated protein kinase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: P45984
Assay: Inhibition of His6-tagged JNK2 (unknown origin) assessed as reduction in c-Jun phosphorylation using GST-c-Jun (1 to 79 residues) and [gamma32P]ATP incubated for 1 hr by scintillation counting assay
Measurement: IC50 <= 1000 nM (pChEMBL 6)
Descriptors: MW 317.35, ALogP 3.49, HBA 3, HBD 2, PSA 81, QED 0.61
Compound (SMILES): N#C[C@@H]1CCN(CC2CCOC2)C[C@H]1n1cc(C(N)=O)c(Nc2ccc(F)cc2)n1
InChIKey: XTYQZCTXIZSIBN-DZNRSUDRSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: Biochemical HTRF Assay: The ability of compounds to inhibit the activity of JAK1, JAK2, JAK3, and Tyk2 was measured using a recombinant purified GST-tagged catalytic domain for each enzyme (Invitrogen JAK1 #M4290, JAK2 #M4290, JAK3 #M4290, Tyk2 #M4290) in an HTRF format 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 μL of 1.11× enzyme and 1.11× substrate in 1× 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 ambient temperature to allow compound binding to equilibrate. After equilibration, 2 μL of 10×ATP in 1× assay buffer was added to initiate the kinase reaction and the plates were shaken and then incubated at ambient temperature for 120 minutes. At the end of the incubation, 20 μL of 2× 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. Plates were shaken and centrifuged and then incubated 60 minutes at ambient temperature and then read on a Perkin Elmer Envision (λex=337 nm, λem=665 and 615 nm, TRF delay time=20 μs). HTRF signal=10,000*665 nm reading/615 nm reading.
Measurement: IC50 = 6 nM (pChEMBL 8.220000267028809)
Descriptors: MW 412.47, ALogP 2.29, HBA 7, HBD 2, PSA 109.2, QED 0.75
Compound (SMILES): O=C(Nc1ccc(C(=O)N2CCN(c3ncccn3)CC2)cc1)N1Cc2ccc(F)cc2C1
InChIKey: PPJADNKOAMVKMG-UHFFFAOYSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Binding Assay : Probe 1:The assay was carried out in 18 uL low volume plates (Owens Corning) in reaction buffer (50 mM HEPES (NaOH), pH 7.5, 100 mM NaCl, 10 mM MgCl2, 1 mM DTT, 1% Glycerol) using 6.8 nM recombinant, human, C-terminally-His tagged NAMPT, 1 nM Tb-anti-His antibody (Invitrogen, Cat #PV5895), and 200 nM probe (Oregon Green 488-conjugated APO0866; A-1251667.0. Plates were covered, and reactions were carried out for 2-3 hours. Plates were read with Envision (Laser Lantha low volume protocol) after 2 to 3 hours. Excitation was carried out at 337 nm, and the ratio of emission of Oregon Green (520 nm) to terbium (492 nm) was determined and used to calculate IC50 values of test compounds.
Measurement: IC50 = 11 nM (pChEMBL 7.960000038146973)
Descriptors: MW 446.49, ALogP 3.13, HBA 5, HBD 1, PSA 81.67, QED 0.67
Compound (SMILES): Cc1ccc(C2=C(NC(=O)c3cc(Br)no3)C(=O)NC(c3ccc(OCCCC(F)(F)F)cc3)(C(F)(F)F)C2)cc1
InChIKey: ITICLGIEMYOFMG-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: MGAT LCMS Assay: The MGAT enzyme reactions were performed in CORNING Falcon 96-well Polypropylene plates, in a total volume of 60 uL of 50 mM Potassium Phosphate buffer pH 7.4, containing a final concentration of 100 uM 2-oleoylglycerol, 15 uM oleoyl-Coenzyme A and 0.0013 ug/uL Human or Mouse MGAT-2 or 0.0026 ug/uL Rat recombinant MGAT-2 membranes expressed in Sf9 cells. Assay plates were run through a fully automated robotics system and shaken for 5 seconds every minute for a total 10 minutes. The reactions were then quenched with 120 uL of ice cold methanol containing 1 ug/mL 1,2-distearoyl-rac-glycerol as the internal standard. Plates were shaken for 2 minutes and spun down to remove protein precipitation. After the spin, samples were transferred to LC/MS compatible PCR plates. For LC/MS analysis, a ThermoFisher Surveyor pump, utilizing a Waters Symmetry C8, 50x2.1 mm column, was used for the chromatography of enzyme products.
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 646.38, ALogP 6.59, HBA 5, HBD 2, PSA 93.46, QED 0.21
Compound (SMILES): CCC(CC)(NC(=O)c1ccc(N2CC(F)(F)C2)c(OCC2CC2)n1)C(=O)N1CC(F)(F)C1
InChIKey: UZHGOWURUWWMQW-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1.times.HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37° C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30.degree. C. for 30 min Compounds were added to a final assay volume of 100 ul and incubated for 30 min at 30° C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 ul lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN.sub.3) and 50 ul detection solutions (20 uM mAb Alexa700-cAMP 1:1, and 48 uM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated.
Measurement: EC50 = 4.7 nM (pChEMBL 8.329999923706055)
Descriptors: MW 472.48, ALogP 3.09, HBA 5, HBD 1, PSA 74.77, QED 0.56
Compound (SMILES): NC(=O)c1c(-c2ccc(Oc3ccccc3)cc2)nn2c1NCCC21CCN(Cc2ccccc2)CC1
InChIKey: FCRQWPYVHQHSFX-UHFFFAOYSA-N
Target: Tyrosine-protein kinase BTK (SINGLE PROTEIN), Homo sapiens
UniProt: Q06187
Assay: Time-Resolved Fluorescence Resonance Energy Transfer Assay: Compounds disclosed herein were tested for inhibition of Btk kinase activity in an assay based on time-resolved fluorescence resonance energy transfer methodology. Recombinant Btk was pre-incubated with the compounds disclosed herein at room temperature for 1 hour in an assay buffer containing 50 mM Tris pH7.4, 10 mM MgCl2, 2 mM MnCl2, 0.1 mM EDTA, 1 mM DT, 20 nM SEB, 0.1% BSA, 0.005% tween-20. The reactions were initiated by the addition of ATP (at the concentration of ATP Km) and peptide substrate (Biotin-AVLESEEELYSSARQ-NH2). After incubating at room temperature for 1 h, an equal volume of stop solution containing 50 mM HEPES pH7.0, 800 mM KF, 20 mM EDTA, 0.1% BSA, Eu cryptate-conjugated p-Tyr66 antibody and streptavidin-labeled XL665 was added to stop the reaction. Plates were further incubated at room temperature for 1 hour, and then the TR-FRET signals (ex337 nm, em 620 nm/665 nm) were read on BMG PHERAstar FS instrument. The residual enzyme activity in presence of increasing concentrations of compounds was calculated based on the ratio of fluorescence at 615 nm to that at 665 nm. The IC50 for each compound was derived from fitting the data to the four-parameter logistic equation by Graphpad Prism software.
Measurement: IC50 = 180 nM (pChEMBL 6.75)
Descriptors: MW 493.61, ALogP 5.25, HBA 6, HBD 2, PSA 85.41, QED 0.38
Compound (SMILES): CC(C)(COc1ccc(-c2ccc(-c3ncc(C4CC(F)(F)C4)[nH]3)cn2)cc1)C(=O)O
InChIKey: LCXFQWQBTMONQC-UHFFFAOYSA-N
Target: Diacylglycerol O-acyltransferase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: O75907
Assay: DGAT1 Inhibitory Assay: As a buffer to be used in the enzymatic reaction of DGAT1, 100 mM Tris-HCl (pH 7.4), 200 mM Sucrose, 20 mM MgCl2, 0.125% Bovine Serum Albumin (BSA) were used. To the buffer were added Test compound with a predetermined concentration as well as 15 μM dioleoylglycerol, 5 μM [14C]-palmitoyl-CoA, 100 μg protein/nth DGAT1 highly expressed-expresSF+® microsome, 0.75% acetone, and 1% dimethylsulfoxide, and triglyceride (TG) synthetic reaction was carried out at 30 °C. for 20 minutes with a volume of 100 μL. 90 μL of the reaction solution was added to 810 μL of methanol to stop the reaction. The reaction solution was added to Oasie® μElution plate (available from Waters Corporation), and eluted with 150 μL of a mixed solution of acetonitrile: isopropanol (=2:3). To elute was added 150 μL of MicroScinti™-40 (available from PerkinElmer Inc.), and after thoroughly stirring the mixture, a [14C]-TG amount formed by the reaction was quantitated by measuring the same using TopCount™-NXT (available from PerkinElmer Inc.).
Measurement: IC50 = 14 nM (pChEMBL 7.849999904632568)
Descriptors: MW 427.45, ALogP 5.14, HBA 4, HBD 2, PSA 88.1, QED 0.55
Compound (SMILES): CN(C)c1nc2c(c(NCc3ccc(C(F)(F)F)nc3)n1)CCN(c1ncccc1C(F)(F)F)CC2
InChIKey: ALQRBYBHEJRSAY-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8NER1
Assay: Functional Assay: HEK293 cells were transfected with human TRPV1 cloned in pcDNA3.1zeo(+) using the Effectene non-liposomal lipid based transfection kit (Qiagen) (hTRPV1/HEK293). hTRPV1/HEK293 cells were routinely grown as monolayers under selection in zeocin (200 μg/mL; Invitrogen) in Dulbecco's Modified Eagle Medium (DMEM, Gibco BRL) supplemented with 10% fetal bovine serum, and penicillin/streptomycin (50 units/mL) 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 = 90 nM (pChEMBL 7.050000190734863)
Descriptors: MW 511.47, ALogP 4.59, HBA 7, HBD 1, PSA 70.07, QED 0.5
Compound (SMILES): NS(=O)(=O)OC[C@@H]1C[C@@H](Nc2ncnc3[nH]c(-c4ccccc4)nc23)C[C@@H]1O
InChIKey: WKOLOFZQDVCYHM-XQQFMLRXSA-N
Target: NEDD8 activating enzyme (PROTEIN COMPLEX), Homo sapiens
UniProt: Q8TBC4
Assay: Inhibition of recombinant human NAEalpha/N-terminal His-tagged NAEbeta expressed in baculovirus infected insect Sf9 cells using N-terminal FLAG tagged Nedd8 as substrate incubated for 105 mins in presence of N-terminal GST-tagged Ubc12 by FRET assay
Measurement: IC50 <= 500 nM (pChEMBL 6.301000118255615)
Descriptors: MW 404.45, ALogP 0.79, HBA 8, HBD 4, PSA 156.11, QED 0.47
Compound (SMILES): NS(=O)(=O)OC[C@@H]1C[C@@H](Nc2ncnc3[nH]c(C4CCCCC4)nc23)C[C@@H]1O
InChIKey: KTFPISCAIDIOGI-XQQFMLRXSA-N
Target: NEDD8 activating enzyme (PROTEIN COMPLEX), Homo sapiens
UniProt: Q8TBC4
Assay: Inhibition of recombinant human NAEalpha/N-terminal His-tagged NAEbeta expressed in baculovirus infected insect Sf9 cells using N-terminal FLAG tagged Nedd8 as substrate incubated for 105 mins in presence of N-terminal GST-tagged Ubc12 by FRET assay
Measurement: IC50 <= 500 nM (pChEMBL 6.301000118255615)
Descriptors: MW 410.5, ALogP 1.17, HBA 8, HBD 4, PSA 156.11, QED 0.55
Compound (SMILES): Cl.O=c1c(CO)cc2cnc(Nc3ccc(N4CCNCC4)cn3)nc2n1C1CCCC1
InChIKey: PBWYPNJVIZCRDR-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 4/cyclin D1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P11802
Assay: Inhibition of Cdk4/Cyclin D (unknown origin) using retinoblastoma protein substrate and [gamma32P]ATP incubated for 15 mins by beta plate counter based assay
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 421.51, ALogP 1.95, HBA 9, HBD 3, PSA 108.2, QED 0.57
Compound (SMILES): N=C(N)NCCCCC(=O)Nc1ccc(C[C@H](NC(=O)[C@@H]2CCCCN2S(=O)(=O)c2ccccc2)C(=O)O)cc1
InChIKey: VTJZZOUBHILYBS-GOTSBHOMSA-N
Target: Integrin alpha-V/beta-1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P05556
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 = 0.3162 nM (pChEMBL 9.5)
Descriptors: MW 572.69, ALogP 1.63, HBA 6, HBD 6, PSA 194.78, QED 0.12
Compound (SMILES): CN1CCN(CC(O)CN2CCc3ccccc3C2)C(=O)C1
InChIKey: SVZKIGMIDKRMFU-UHFFFAOYSA-N
Target: PRMT5/MEP50 complex (PROTEIN COMPLEX), Homo sapiens
UniProt: O14744
Assay: Inhibition of recombinant human fetal brain FLAG-tagged PRMT5/full length human testis 6His-tagged MEP50 expressed in baculovirus infected insect Sf9 cells using human histone H4 (1 to 15 residues) as substrate preincubated for 30 mins followed by [3H]-SAM addition
Measurement: IC50 > 50000 nM (pChEMBL 4.301000118255615)
Descriptors: MW 303.41, ALogP 0.18, HBA 4, HBD 1, PSA 47.02, QED 0.86
Compound (SMILES): O=C(Nc1ccc(CNc2nc(N3CCCC3)nc3ccc(C(F)(F)F)cc23)cc1)C1CCN(Cc2ccc(F)cc2)CC1
InChIKey: DTHMTMSFHVHDNE-UHFFFAOYSA-N
Target: TCF4/beta-catenin (PROTEIN COMPLEX), Homo sapiens
UniProt: Q9NQB0
Assay: Inhibition of TCF-4/beta catenin in human Tcf33.13 cells expressing Tcf-4/SV40 binding sites driving firefly luciferase reporter gene after 24 hrs by luminometer
Measurement: IC50 = 3691 nM (pChEMBL 5.429999828338623)
Descriptors: MW 606.67, ALogP 6.85, HBA 6, HBD 2, PSA 73.39, QED 0.21
Compound (SMILES): CN1CCC[C@H]1COc1cc(O)cc(Br)c1
InChIKey: QTXUGSOGZYIQNZ-JTQLQIEISA-N
Target: Neuronal acetylcholine receptor; alpha3/beta4 (PROTEIN COMPLEX), Homo sapiens
UniProt: P30926
Assay: Displacement of [3H]-epibatine from human alpha3beta4-nACHR expressed in HEK243 cell membranes preincubated for 5 mins followed by overnight incubation with [3H]-epibatine by liquid scintillation counting analysis
Measurement: Ki = 147 nM (pChEMBL 6.829999923706055)
Descriptors: MW 286.17, ALogP 2.63, HBA 3, HBD 1, PSA 32.7, QED 0.93
Compound (SMILES): O=S(=O)(Nc1ccc(-c2nc3ccc(NS(=O)(=O)c4ccc5ccccc5c4)cc3[nH]2)cc1)c1ccc2ccccc2c1
InChIKey: SCVWSOAMWJCBHO-UHFFFAOYSA-N
Target: GroEL/GroES (PROTEIN COMPLEX), Escherichia coli
UniProt: Q548M1
Assay: Inhibition of Escherichia coli GroEL/GroES expressed in Escherichia coli DH5alpha/BL21 (DE3) assessed as inhibition of denatured MDH refolding preincubated for 10 mins followed by ATP addition measured after 20 to 40 mins by fluorescence based assay
Measurement: IC50 = 40000 nM (pChEMBL 4.400000095367432)
Descriptors: MW 604.71, ALogP 7.14, HBA 5, HBD 3, PSA 121.02, QED 0.18
Compound (SMILES): O=C(Cc1ccccc1)NC[C@H](O)CN1CCc2ccccc2C1
InChIKey: IVYDHNJIVLFCFB-IBGZPJMESA-N
Target: PRMT5/MEP50 complex (PROTEIN COMPLEX), Homo sapiens
UniProt: O14744
Assay: Inhibition of recombinant human fetal brain FLAG-tagged PRMT5/full length human testis 6His-tagged MEP50 expressed in baculovirus infected insect Sf9 cells using human histone H4 (1 to 15 residues) as substrate preincubated for 30 mins followed by [3H]-SAM addition
Measurement: IC50 = 101 nM (pChEMBL 7)
Descriptors: MW 324.42, ALogP 1.76, HBA 3, HBD 2, PSA 52.57, QED 0.85
Compound (SMILES): CNc1nc(NCc2ccc(NC(=O)c3ccc(OC(F)(F)F)cc3)cc2)c2ccccc2n1
InChIKey: SPLWHRKCICYLBP-UHFFFAOYSA-N
Target: TCF4/beta-catenin (PROTEIN COMPLEX), Homo sapiens
UniProt: Q9NQB0
Assay: Inhibition of TCF-4/beta catenin in human Tcf33.13 cells expressing Tcf-4/SV40 binding sites driving firefly luciferase reporter gene after 24 hrs by luminometer
Measurement: IC50 = 1232 nM (pChEMBL 5.909999847412109)
Descriptors: MW 467.45, ALogP 5.43, HBA 6, HBD 3, PSA 88.17, QED 0.34
Compound (SMILES): O=C(c1cncc(-c2cccc(OCC(O)CN3CCc4ccccc4C3)c2)c1)N1CCCC1
InChIKey: PAFUFJGRVFVNJB-UHFFFAOYSA-N
Target: PRMT5/MEP50 complex (PROTEIN COMPLEX), Homo sapiens
UniProt: O14744
Assay: Inhibition of recombinant human fetal brain FLAG-tagged PRMT5/full length human testis 6His-tagged MEP50 expressed in baculovirus infected insect Sf9 cells using human histone H4 (1 to 15 residues) as substrate preincubated for 30 mins followed by [3H]-SAM addition
Measurement: IC50 = 1001 nM (pChEMBL 6)
Descriptors: MW 457.57, ALogP 3.78, HBA 5, HBD 1, PSA 65.9, QED 0.58