lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): NC(=O)c1ccc2[nH]c(-c3ccc(OC4CCCN(Cc5ccccc5)C4)cc3)nc2c1
InChIKey: HXLLSHXMSSCPHS-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase Chk2 (SINGLE PROTEIN), Homo sapiens
UniProt: O96017
Assay: Inhibition of human CDS1 using N-terminal biotin labeled SGLYRSPSMPENLNRPR as substrate in presence of [gamma-33P]-ATP after 3 hrs by scintillation counting method
Measurement: IC50 = 63 nM (pChEMBL 7.199999809265137)
Descriptors: MW 426.52, ALogP 4.37, HBA 4, HBD 2, PSA 84.24, QED 0.48
Compound (SMILES): Cc1cncc(-c2cc(Nc3nn(C)cc3C(F)(F)F)nc3[nH]cc(C#N)c23)c1
InChIKey: GRGHLILILXVPIC-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: Homogeneous Time Resolved-fluorescence Resonance Energy Transfer (TR-FRET) Assay: LRRK2 kinase activity is measured using a LanthaScreen kinase activity assay available from Invitrogen (Life Technologies Corporation). The assay is a homogeneous time resolved-fluorescence resonance energy transfer (TR-FRET) assay that measures phosphorylation of a fluorescein-labelled peptide substrate (fluorescein-LRRKtide, fluorescein-GAGRLGRDKYKTLRQIRQ) (SEQ ID NO 1) as a result of LRRK2 kinase activity. The phosphorylated peptide is recognized by a terbium-labelled phospho-specific anti-LRRKtide antibody and, subsequently, the phosphorylated LRRKtide can be quantified by the extent of TR-FRET between the terbium donor and fluorescein acceptor.The LRRK2 kinase is obtained from Invitrogen (Life Technologies Corporation) and comprises residues 970 to 2527 of the full length human wild-type LRRK2 kinase or a similar sequence with the G2019S mutation. As discussed above, this mutation increases the kinase activity relative to the wild-type. The kinase reactions are performed in a 20 uL volume in 384-well plates. The kinase reaction buffer consists of 50 mM Tris pH 8.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA, and 2 mM DTT.Â  In the assay, 1 nM LRRK2 WT or 250 pM LRRK2 G2019S kinase is incubated with the test compound (typically at 0 to 30 uM) for 30 minutes before the kinase reaction is initiated by addition of 1.3 mM ATP and 0.4 uM fluorescein-LRRKtide. The reaction mixture (20 ul total volume) is incubated for 2 hours at 30° C. before the reaction is terminated by addition of 10 mM EDTA and 1 nM terbium-labelled anti-phospho-LRRKtide antibody (final volume 20 ul). The mixture is further incubated for 30 minutes at RT. TR-FRET is measured by excitation of the terbium-donor with 340 nm light and subsequent (delay time 100 us) measurement of terbium and fluorescein emission at 495 nm and 520 nm, respectively, over a time window of 1000 us. The measurement is repeated 10 times for fluorescein and 10 times for terbium emission with a 2000 us time window between repeats. TR-FRET measurements are performed on a Biomek Synergy plate. The TR-FRET signal is calculated as the emission-ratio at 520 nm over 495 nm.
Measurement: IC50 = 509 nM (pChEMBL 6.289999961853027)
Descriptors: MW 397.36, ALogP 4.3, HBA 6, HBD 2, PSA 95.21, QED 0.54
Compound (SMILES): Cn1ccc(Nc2cc(-c3ccccc3)c3c(C#N)c[nH]c3n2)n1
InChIKey: XXZAJGJUAVDOEK-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: Homogeneous Time Resolved-fluorescence Resonance Energy Transfer (TR-FRET) Assay: LRRK2 kinase activity is measured using a LanthaScreen kinase activity assay available from Invitrogen (Life Technologies Corporation). The assay is a homogeneous time resolved-fluorescence resonance energy transfer (TR-FRET) assay that measures phosphorylation of a fluorescein-labelled peptide substrate (fluorescein-LRRKtide, fluorescein-GAGRLGRDKYKTLRQIRQ) (SEQ ID NO 1) as a result of LRRK2 kinase activity. The phosphorylated peptide is recognized by a terbium-labelled phospho-specific anti-LRRKtide antibody and, subsequently, the phosphorylated LRRKtide can be quantified by the extent of TR-FRET between the terbium donor and fluorescein acceptor.The LRRK2 kinase is obtained from Invitrogen (Life Technologies Corporation) and comprises residues 970 to 2527 of the full length human wild-type LRRK2 kinase or a similar sequence with the G2019S mutation. As discussed above, this mutation increases the kinase activity relative to the wild-type. The kinase reactions are performed in a 20 uL volume in 384-well plates. The kinase reaction buffer consists of 50 mM Tris pH 8.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA, and 2 mM DTT.Â  In the assay, 1 nM LRRK2 WT or 250 pM LRRK2 G2019S kinase is incubated with the test compound (typically at 0 to 30 uM) for 30 minutes before the kinase reaction is initiated by addition of 1.3 mM ATP and 0.4 uM fluorescein-LRRKtide. The reaction mixture (20 ul total volume) is incubated for 2 hours at 30° C. before the reaction is terminated by addition of 10 mM EDTA and 1 nM terbium-labelled anti-phospho-LRRKtide antibody (final volume 20 ul). The mixture is further incubated for 30 minutes at RT. TR-FRET is measured by excitation of the terbium-donor with 340 nm light and subsequent (delay time 100 us) measurement of terbium and fluorescein emission at 495 nm and 520 nm, respectively, over a time window of 1000 us. The measurement is repeated 10 times for fluorescein and 10 times for terbium emission with a 2000 us time window between repeats. TR-FRET measurements are performed on a Biomek Synergy plate. The TR-FRET signal is calculated as the emission-ratio at 520 nm over 495 nm.
Measurement: IC50 = 58 nM (pChEMBL 7.239999771118164)
Descriptors: MW 314.35, ALogP 3.58, HBA 5, HBD 2, PSA 82.32, QED 0.61
Compound (SMILES): C[C@]12CC[C@H]3[C@@H](CCC4=CC(=O)CC[C@@]43C)C1=CC=C2n1ccnc1
InChIKey: RTRGJRBCXDZOOW-UDDWMFPDSA-N
Target: Steroid 17-alpha-hydroxylase/17,20 lyase (SINGLE PROTEIN), Homo sapiens
UniProt: P05093
Assay: Inhibition of CYP17 in human testicular microsomes using 17alpha-hydroxypregnenolone as substrate assessed as decrease in DHEA levels after 90 mins by ELISA
Measurement: IC50 <= 11 nM (pChEMBL 7.959000110626221)
Descriptors: MW 334.46, ALogP 4.79, HBA 3, HBD 0, PSA 34.89, QED 0.74
Compound (SMILES): COc1ccccc1C(=O)Nc1cccc(CN2CCC(C(=O)NC3CCCCC3)CC2)c1
InChIKey: GORZFWDCYFBLGD-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 = 17 nM (pChEMBL 7.769999980926514)
Descriptors: MW 449.6, ALogP 4.61, HBA 4, HBD 2, PSA 70.67, QED 0.65
Compound (SMILES): COC1CCCN(c2cc(-c3n[nH]c4ccc(OC(C)C)cc34)ncn2)C1
InChIKey: IXEGROFEYMCIIY-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: LanthaScreen™ Assay (5 mM ATP): a) 400 nl of a 1:2.15 serial dilution of test compound(98 μM top assay concentration) is spotted via Labcyte Echo to certain wells in a 384 well black, untreated plate. Control wells contain 400 nl of either DMSO or 400 nl of a known inhibitor in DMSO. b) 10 μl of a 2.5 nM LRRK2(G2019S mutation, GST-LRRK2(amino acids 970-2527)) enzyme solution in 1× assay buffer(50 mM Tris pH 8.5, 10 mM MgCl2, 0.01% Brij-35, 1.0 mM EGTA, 2 mM DTT, 0.05 mM NaVO4) is added to all wells. c) A 30 minute room temperature incubation is followed by addition of 10 μl of 800 nM fluorescein labeled LRRKtide peptide substrate and 10 mM ATP solution in 1× assay buffer to all wells. d) After a 35 minute room temperature incubation, 20 μl of TR-FRET Dilution Buffer(Invitrogen PV3756B) containing 4 nM Tb-labeled anti-phospho LRRKtide antibody and 20 mM EDTA is added to all wells. e) Plates are incubated at room temperature for 1 hour and read on an Envision™ multi-mode plate reader with LanthaScreen™ settings. Results are analysed using Assay Data Analyzer.
Measurement: IC50 = 12.7 nM (pChEMBL 7.900000095367432)
Descriptors: MW 367.45, ALogP 3.42, HBA 6, HBD 1, PSA 76.16, QED 0.74
Compound (SMILES): O=C(CCCCCCCNC(=O)c1nc(-c2nccs2)sc1C1CC1)NO
InChIKey: JFVGWEKFOOWMDP-UHFFFAOYSA-N
Target: Histone deacetylase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13547
Assay: Enzyme Assay: Enzyme activity of HDAC1,3 is determined using the substrate Ac-Lys-Tyr-Lys(Ac)-AMC while the enzyme activity of HDAC6 is assayed using the substrate Boc-lys(Ac)-AMC. The reaction is carried out in flat-bottom, 96-well, or 384-well microplates. After the substrate was deacetylated by HDACs, the product AMC which was obtained from hydrolysis by trypsin can generate fluorescence signal. Measurement is taken in using an multilabe plate reader a 355 nm excitation filter and a 460 nm emission filter. The initial rate of fluorescence should accurately reflect the rate of product formation and enzyme activity. Sample processing: The sample was dissolved in DMSO and kept at a low temperature. DMSO in the final system is limited under a low concentration which won't affect the enzyme activity.
Measurement: IC50 = 86 nM (pChEMBL 7.070000171661377)
Descriptors: MW 408.55, ALogP 3.72, HBA 7, HBD 3, PSA 104.21, QED 0.3
Compound (SMILES): CC1(C)CCC(CN2CCN(c3ccc(C(=O)NS(=O)(=O)c4ccc(NCC5CCOCC5)c([N+](=O)[O-])c4)c(Oc4cccc5c4CCNC5)c3)CC2)=C(c2ccc(Cl)cc2)C1
InChIKey: CBBCCUNTGPYKBL-UHFFFAOYSA-N
Target: Apoptosis regulator Bcl-2 (SINGLE PROTEIN), Mus musculus
UniProt: P10417
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 = 2.923 nM (pChEMBL 8.529999732971191)
Descriptors: MW 883.51, ALogP 8.43, HBA 11, HBD 3, PSA 155.38, QED 0.09
Compound (SMILES): COc1cc(-c2cc(F)cnc2OC)ccc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: BTLMGPIWTWZTDZ-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 = 102 nM (pChEMBL 6.989999771118164)
Descriptors: MW 522.51, ALogP 4, HBA 9, HBD 1, PSA 125.55, QED 0.34
Compound (SMILES): O=C(O)COc1ccc(Cl)cc1[C@@H]1c2scnc2CCN1C(=O)OCc1cccc(F)c1F
InChIKey: PPGSXGUNCMXHHT-HXUWFJFHSA-N
Target: Prostaglandin D2 receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Y4
Assay: Radioligand Displacement Assay: Binding assay was performed in a final assay volume of 250 μl. First, 25 μl of test compound, previously diluted in Binding-Buffer (Binding-Buffer: 50 mM Tris-Base, 100 mM NaCl, 1 mM EDTA, 0.1% BSA (protease free), 0.01% NaN3, 10 mM MnCl2, pH 7.0) was placed into each well. After addition of 75 μl Binding-Buffer, 50 μl of the radioligand 3H-PGD2 (at 2.5 nM (220.000 dpm/well) from ANAWA ART0662) was added to each well. Binding assay was started by addition of 100 μl CRTH2 membrane fragments, reaching a final concentration of 20 μg/well. For non-specific binding, PGD2 was added to the reaction mixture to 10 mM final concentration. This assay mix was incubated for 90 minutes at room temperature and then filtered through a GF/C filter 96-well plate which was pre-soaked for 3 hours in 0.5% polyethyleneimine (PEI). The filter-wells were washed three times with ice cold Binding-Buffer. Then, 40 μl of Microscint-40 (Packard) was added to each well and the re
Measurement: IC50 = 0.5 nM (pChEMBL 9.300000190734863)
Descriptors: MW 494.9, ALogP 4.82, HBA 6, HBD 1, PSA 88.96, QED 0.53
Compound (SMILES): N[C@H](C(=O)O)C(NS(=O)(=O)c1ccc(-c2ccccc2)cc1)C(=O)O
InChIKey: PDTYFUFTPPYALH-LSLKUGRBSA-N
Target: Excitatory amino acid transporter 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P43005
Assay: Inhibition of [3H]-D-Asp uptake at human EAAT3 expressed in HEK293 cells measured after 3 mins by TopCount method
Measurement: IC50 = 20417.38 nM (pChEMBL 4.690000057220459)
Descriptors: MW 364.38, ALogP 0.5, HBA 5, HBD 4, PSA 146.79, QED 0.56
Compound (SMILES): C/C(=C\c1cc(F)c(Oc2ccc(S(=O)(=O)NCCOCCOCCOCCOCCOCCOCCNC(=O)C(O)C(O)C(=O)NCCOCCOCCOCCOCCOCCOCCNS(=O)(=O)c3ccc(Oc4c(F)cc(/C=C(\C)C(=O)N=C(N)N)cc4F)cc3)cc2)c(F)c1)C(=O)N=C(N)N
InChIKey: ZNMHSIDKSDXRIU-JUWAPXIKSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P26433
Assay: Inhibition of rat NHE-3-mediated Na+ dependent H+ antiport expressed in opossum kidney cells in presence of NHE-1 antagonist EIPA by BCECF-AM dye based fluorescence assay
Measurement: IC50 = 1000 nM (pChEMBL 6)
Descriptors: MW 1549.63
Compound (SMILES): COc1ncccc1C(=O)Nc1cccc(CN2CCC(C(=O)NC3CCCCC3)CC2)c1
InChIKey: HYZQLGRUYQXBKY-UHFFFAOYSA-N
Target: Atypical chemokine receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P25106
Assay: In Vitro Assay: CHO-K1 CXCR7 b-arrestin cells are detached from culture dishes with a cell dissociation buffer (Invitrogen, #13151-014) and collected in growing medium (F12 HAMS 90% (v/v)/FCS 10% (v/v), Penicilin/streptomycin 1% (v/v)). 5000 cells per well (in 20 μl) are seeded in a 384 well plate (white-walled, clear bottom; BD Falcon #353274). The plate is incubated at 37° C./5% CO2 for 24 hours. Medium is then replaced by 20 μl OPTIMEM (Invitrogen #31985) for 3 to 4 hours. Test compounds are dissolved at 10 mM in DMSO and serially diluted in DMSO to 200× of the final concentration for dose response curves. Compounds are then diluted 1:33.3 in HBSS1×. 5 μl/well of HBSS1×/20 mM HEPES/0.2% BSA are added to the assay plate followed by addition of 5 μl/well of diluted compounds. CXCL12 (Peprotech #300-28A) may be used as a reference agonist. The plate is incubated for 90 minutes at 37° C. 12 μl of detection reagent (Path Hunter Detection Kit, DiscoveRx, #93-0001) is transferred to the assay plate and to the plate is incubated for 1 hour at room temperature. Luminescent signal is read in a microplate reader (FLUOstar Optima, bmg).
Measurement: EC50 = 10 nM (pChEMBL 8)
Descriptors: MW 450.58, ALogP 4, HBA 5, HBD 2, PSA 83.56, QED 0.67
Compound (SMILES): CN1C(=O)N(c2cccc(C(F)(F)F)c2)C2=C(C(=O)CC2)C1c1ccc(C#N)cc1C(=O)NC1COC1
InChIKey: MTEYTVRTZYWDIA-UHFFFAOYSA-N
Target: Neutrophil elastase (SINGLE PROTEIN), Homo sapiens
UniProt: P08246
Assay: Human Neutrophil Elastase Assay: 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 μL of these compound dilutions were mixed with 10 μl 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 μL substrate solution in assay buffer were added (250 μM 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.
Measurement: IC50 = 5.3 nM (pChEMBL 8.279999732971191)
Descriptors: MW 510.47, ALogP 3.94, HBA 5, HBD 1, PSA 102.74, QED 0.67
Compound (SMILES): Cc1nc(C(C)(CS(C)(=O)=O)NC(=O)c2cc(OCC(F)(F)F)c(C3(F)COC3)cn2)no1
InChIKey: PRYDBTFZRIASDS-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×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 = 333.4 nM (pChEMBL 6.480000019073486)
Descriptors: MW 496.44, ALogP 1.6, HBA 9, HBD 1, PSA 133.51, QED 0.54
Compound (SMILES): CC(F)(F)c1cncc(NC(=O)Nc2ccc(Oc3ccnc(NC(=O)C4CC4)c3)c(F)c2)c1
InChIKey: OQJZBWFPSBLNRC-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 = 17.5 nM (pChEMBL 7.760000228881836)
Descriptors: MW 471.44, ALogP 5.51, HBA 5, HBD 3, PSA 105.24, QED 0.42
Compound (SMILES): CCS(=O)(=O)N1CC(CC#N)(n2cc(C(N)=O)c(Nc3ccc(C(=O)N4CCS(=O)(=O)CC4)cc3)n2)C1
InChIKey: DFQUTVGNAWWAQW-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
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 = 27 nM (pChEMBL 7.570000171661377)
Descriptors: MW 549.64, ALogP -0.13, HBA 10, HBD 2, PSA 188.56, QED 0.45
Compound (SMILES): CCC1CC(=O)NN=C1c1ccc2nc(-c3cccc(OCCN(C(C)C)C(C)C)c3)oc2c1
InChIKey: LPVRPHLCDIZAAM-UHFFFAOYSA-N
Target: Glucose-dependent insulinotropic receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q8TDV5
Assay: AlphaScreen cAMP Assay: MIN6 cells [Miyazaki J et al. Endocrinology. 1990 July; 127(1):126-32] were stably transfected with an expression vector for human GPR119 cDNA (Acc. No. NP-848566). Min-6/hGPR119 cells are cultured in DMEM, 10% FBS, 50 uM beta-mercaptoethanol, 0.3 mg/mL Geniticin, 2 mM GlutaMAX at 37 C. 5% CO2. For the assay, the cells are placed in Optiplates (white, 384-well, 160W-barcoded, TC, sterile with lid, Cat. No. #6007688 (Perkin Elmer); 10000 cells/well; 50 ul). The plates covered with lids are then incubated for 24 hours at 37 C./5% CO2.After the medium has been sucked out of the wells completely, 10 ul of the test compound are added, the compounds are diluted beforehand with stimulating buffer (140 mM NaCl, 3.6 mM KCl, 0.5 mM NaH2PO4, 0.5 mM MgSO4, 1.5 mM CaCl2, 10 mM Hepes, 5 mM NaHCO3; pH 7.4. 0.5 mM IBMX and 0.1% BSA, the final DMSO concentration is 1%). After 45 minutes' incubation at RT, the cAMP concentrations are determined using the AlphaScreen cAMP Assay.
Measurement: EC50 = 424 nM (pChEMBL 6.369999885559082)
Descriptors: MW 462.59, ALogP 5.24, HBA 6, HBD 1, PSA 79.96, QED 0.47
Compound (SMILES): O=C(c1ccc(-c2cccs2)nc1)N1CCC(N2C(=O)OCc3ccccc32)CC1
InChIKey: GQPVADGJKGUOEG-UHFFFAOYSA-N
Target: Hematopoietic prostaglandin D synthase (SINGLE PROTEIN), Homo sapiens
UniProt: O60760
Assay: Biochemical Assay: The H-PGDS catalyzed conjugation of GSH and 1-chloro-2,4-dinitrobenzene (CDNB) was used as the biochemical assay for enzyme inhibition. Reactions were performed in 96 well plate format, and product formation was followed at A340 nm over a 10 min interval at 25° C. using a POWERWAVE XS microplate scanning spectrophotometer (Bio-Tek Instruments, Winooski, Vt., USA). Reactions were performed in 0.1 M Tris HCl, pH 8.0 containing 2 mM MgCl2, 1 mM CDNB, 2 mM GSH, 2.5 ng/μ1 purified H-PGDS and 10% (v/v) ethanol in a 200 μl reaction volume. IC50 values were calculated from rates of conjugation activity determined at eight concentration points bracketing the IC50, where compound solubility allowed, and were corrected for background activity at the same solvent concentrations. All compounds were made up in 100% DMSO and diluted with 0.1 M Tris HCl, pH 8.0 with 2 mM MgCl2.
Measurement: IC50 = 3400 nM (pChEMBL 5.46999979019165)
Descriptors: MW 419.51, ALogP 4.57, HBA 5, HBD 0, PSA 62.74, QED 0.62
Compound (SMILES): C[C@@H]1CN([C@@H](C)CO)S(=O)(=O)c2ccc(/C=C/c3ccccc3)cc2O[C@H]1CN(C)C(=O)NC1CCCCC1
InChIKey: DIBJIDYNEHVBAV-KFRKLSNLSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Inhibition of human C-terminal His8-tagged IDH1-R132H mutant expressed in Escherichia coli Rosetta assessed as inhibition of ketoreductase activity by measuring NADPH consumption preincubated for 60 mins followed by addition of alpha-KG as substrate measured after 50 mins in presence of NADPH and Mg2+ by resazurin based diaphorase-coupled fluorescence assay
Measurement: IC50 = 190 nM (pChEMBL 6.71999979019165)
Descriptors: MW 555.74, ALogP 4.6, HBA 5, HBD 2, PSA 99.18, QED 0.49
Compound (SMILES): COc1cc(F)ccc1Oc1nc2ccccc2nc1C(=O)Nc1ccc2[nH]c(=O)[nH]c2c1
InChIKey: ZQBFNVOMTKSWEN-UHFFFAOYSA-N
Target: Sodium channel protein type 10 subunit alpha (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q62968
Assay: Electrophysiology Assay: Patch clamp electrophysiology was used to assess the efficacy and selectivity of sodium channel blockers in dorsal root ganglion neurons. Rat neurons were isolated from the dorsal root ganglions and maintained in culture for 2 to 10 days in the presence of NGF (50 ng/ml) (culture media consisted of NeurobasalA supplemented with B27, glutamine and antibiotics). Small diameter neurons (nociceptors, 8-12 um in diameter) were visually identified and probed with fine tip glass electrodes connected to an amplifier (Axon Instruments). The "voltage clamp" mode was used to assess the compound's IC50 holding the cells at -60 mV. In addition, the "current clamp" mode was employed to test the efficacy of the compounds in blocking action potential generation in response to current injections.
Measurement: IC50 = 19 nM (pChEMBL 7.71999979019165)
Descriptors: MW 445.41, ALogP 3.99, HBA 6, HBD 3, PSA 121.99, QED 0.38
Compound (SMILES): COc1ccc(/C=C2\CN(C(=O)Cn3cnnn3)Cc3c(C(=O)N[C@H](C)c4ccccc4)nn(-c4ccc(Cl)cc4Cl)c32)cc1
InChIKey: FYIZKKBMENMFBG-RXULLKCLSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: Inverse agonist activity at human CB2 receptor transfected in HEK293 cells assessed as increase of forskolin-induced cAMP production incubated for 30 mins followed by d2 labeled cAMP addition measured after 60 mins by HTRF assay
Measurement: EC50 = 1130 nM (pChEMBL 5.949999809265137)
Descriptors: MW 643.53, ALogP 5.25, HBA 9, HBD 1, PSA 120.06, QED 0.25
Compound (SMILES): O=C(NCc1ccc(OC(F)(F)F)c(F)c1)C1COc2ncccc21
InChIKey: XCXPKEUZIQGOOP-UHFFFAOYSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: YO-PRO Assay: HEK cells stably expressing P2X7 were used to screen drugs using the YO-PRO1 assay. Cells were grown to confluency in adherent culture at 37° C. in a humidified 5% CO2 incubator (split ⅕ every 3-4 days with DMEM, 10% FCS, 1% Penicillin/Streptomycin, 250 μg/ml Geneticin). Adherent cells were detached by incubation with Trypsine (1 ml per 165 cm2 dish) for 2 minutes, then washed off with 10 ml PBS (without Mg2+ and Ca2+), and resuspended in DMEM, 10% FCS, 1% Penicillin/Streptomycin, no Geneticin. 10,000 cells per well (48 hours before the assay) or 25,000 cells per well (Vi-cell XR (Beckman Coulter) (24 hours before the assay) in 50 μl full medium were seeded on 384-well black-wall, clear bottom plates, that were coated before with 10 μl per well Poly-L-Lysine, incubated for 30-60 minutes at 37° C. and washed once with PBS. Medium was removed from cells and 50 μl of assay buffer containing 0.5 μM YO-PRO-1 was added into the wells. Solutions of antagonist compounds were prepared by serial dilutions of a 10 mM DMSO solution of the antagonist into PBS using a BioMek (Beckman Coulter). Each concentration was performed in duplicate. For IC50 measurements 10 concentration points were measured (10 μM being the highest concentration followed by 9 serial dilution steps ⅓). The cells were incubated with the antagonists of the present invention together with ATP at a final concentration of 250 μM for 90 minutes. During this time period, four time points were taken. Each time point comprised the average of several measurements made within a few seconds. Fluorescence was measured in the FLIPR tetra (Molecular Devices) using the filters appropriate for YO-PRO-1 fluorescence (excitation485/20, emission 530/25).
Measurement: IC50 = 3065 nM (pChEMBL 5.510000228881836)
Descriptors: MW 356.28, ALogP 2.91, HBA 4, HBD 1, PSA 60.45, QED 0.86
Compound (SMILES): O=c1oc2c(O)c(O)ccc2cc1-c1ccc(O)c(O)c1
InChIKey: MZSZXDSVFQKLPB-UHFFFAOYSA-N
Target: Adenosine receptor A1 (SINGLE PROTEIN), Homo sapiens
UniProt: P30542
Assay: Displacement of [3H]CCPA from human adenosine receptor A1 expressed in CHO cell membranes by radioligand competition assay
Measurement: Ki = 6280 nM (pChEMBL 5.199999809265137)
Descriptors: MW 286.24, ALogP 2.28, HBA 6, HBD 4, PSA 111.13, QED 0.4
Compound (SMILES): O=C(Nc1ccc(CNc2nc(NC3CCCCC3)nc3ccccc23)cc1)c1ccc(F)cc1
InChIKey: WLIFRFDORDZAFM-UHFFFAOYSA-N
Target: TCF4/beta-catenin (PROTEIN COMPLEX), Homo sapiens
UniProt: P35222
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 = 364 nM (pChEMBL 6.440000057220459)
Descriptors: MW 469.56, ALogP 6.38, HBA 5, HBD 3, PSA 78.94, QED 0.29
Compound (SMILES): CN1CCC(N2CCN(c3ccc(-c4ccc5[nH]c6nccc(-c7ccc(Cc8ccccc8)cc7)c6c5c4)cc3)CC2)CC1
InChIKey: FHDTVALVKGKOLC-UHFFFAOYSA-N
Target: ALK tyrosine kinase receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UM73
Assay: ELISA-Based In Vitro Kinase Assay: GST-tagged recombinant wild-type or L1196M mutated ALK kinase (rALK) was expressed in Sf9 insect cells using the pBacPAK baculovirus vector system (Clontech) and purified using Glutathione Sepharose 4B affinity beads (GE Healthcare). Recombinant 3C protease was used to remove the GST tag. Purified ALK was used to screen inhibitors in the ELISA-based kinase assay, as follows: Nunc-Immuno 96-well plates were incubated overnight at 30° C. with coating solution containing 2 μg of a specific ALK peptide substrate (ARDIYRASFFRKGGCAMLPVK) in PBS. Wells were then washed with 200 μL of wash buffer (PBS-Tween 0.05%) and incubated with 4% BSA in PBS for at least 2 h at 30° C. The kinase reaction was performed in the presence of 50 mM Tris pH 7.5, 5 mM MnCl2, 5 mM MgCl2, 0.3 mM ATP and purified rALK in a total volume of 100 μL/well at 30° C. for 15 min. For inhibitor testing the reaction mix was preincubated with inhibitor or vehicle for 10 min at room temperature before transferring to the ELISA plate. After the reaction, the wells were washed 5 times with 200 μL of wash buffer. Phosphorylated peptide was detected using 100 μL/well of a mouse monoclonal anti-phosphotyrosine antibody (clone 4G10 UpstateBiotech Ltd) diluted 1:2000 in PBS+4% BSA. After 30 min incubation at room temperature the antibody was removed and wells were washed as described above. 100 μL of a secondary antibody (anti-mouse IgG, Horseradish Peroxidase linked whole antibody, Amersham Pharmacia Biotech) diluted 1:1000 in PBS+4% BSA was added to each well and the plate was incubated again for 30 min at room temperature before washing as above. The plate was developed using 100 μL/well TMB Substrate Solution (Pierce) and the reaction was stopped by adding an equal volume of 1M H2SO4. Finally, the absorbance was read at 450 nm using an ELISA plate reader (Bio-Rad).
Measurement: IC50 = 1500 nM (pChEMBL 5.820000171661377)
Descriptors: MW 591.8, ALogP 7.86, HBA 4, HBD 1, PSA 38.4, QED 0.21
Compound (SMILES): CC1CN(C(=O)c2cc(C#N)ccc2C2C3=C(CCC3=O)N(c3cccc(C(F)(F)F)c3)C(=O)N2C)CCN1
InChIKey: LVRWHOMHRYPQFD-UHFFFAOYSA-N
Target: Neutrophil elastase (SINGLE PROTEIN), Homo sapiens
UniProt: P08246
Assay: Human Neutrophil Elastase Assay: 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 μL of these compound dilutions were mixed with 10 μl 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 μL substrate solution in assay buffer were added (250 μM 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.
Measurement: IC50 = 10.8 nM (pChEMBL 7.96999979019165)
Descriptors: MW 537.54, ALogP 4.24, HBA 5, HBD 1, PSA 96.75, QED 0.63
Compound (SMILES): O=C(N[C@H]1CC[C@@H](O)CC1)c1cc(NC(=O)c2cc(-c3ccc(F)cn3)c(Cl)cc2Cl)n(-c2ccccc2)n1
InChIKey: SMJITQLMIQDIPJ-IZAXUBKRSA-N
Target: High affinity nerve growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P04629
Assay: Cell Based Assay: TRKA co-expressed with p75: The assays are based upon DiscoveRx's proprietary Enzyme Fragment Complementation (EFC) technology. In the case of the TRK cell lines, the enzyme acceptor (EA) protein is fused to a SH2 protein and the TRK receptor of interest has been tagged with a Prolink tag. Cryopreserved PathHunter cells are used from either in-house produced batches or bulk batches bought directly from DiscoveRx. Cryopreserved cells are resuscitated, spun 1000 rpm for 4 min to remove freezing media, and resuspended in MEM+0.5% horse serum (both Invitrogen) to 5e^5 cells/ml. The cells are then plated using a Multidrop into Greiner white tissue culture treated plates at 20 μl/well and incubated for 24 h at 37 °C., 5% CO2, high humidity. On the day of the assay, the cell plates are allowed to cool to room temperature for 30 min prior to the assay. 4 mM stock solutions of test compounds are prepared and serially diluted in 100% DMSO. A standard curve using the compound of Example 135, WO2005/116035 at a top concentration of 150 μM is also prepared on each test plate. High percentage effect (HPE) is defined by 150 μM of the compound of Example 135, WO2005/116035 and 0% effect (ZPE) is defined by 100% DMSO. Plates containing 1 μl of serially diluted compound, standard and HPE/ZPE are diluted 1/66 in assay buffer (PBS minus Ca2+, minus Mg2+ with 0.05% pluronic F127) using a Wellmate. Using a Platemate Plus, 5 μl of 1/66 diluted test compounds is then transferred to the cell plate and allowed to reach equilibrium by incubating for 30 min at room temperature before addition of agonist stimulus: 10 μl/well of 2 nM (0.571 nM FAC) of the cognate neurotrophin (Peprotech) diluted in agonist buffer (HBSS with 0.25% BSA). Final assay concentration of the test compounds is 8.66 μM, (the compound of Example 135, WO2005/116035 FAC is 0.325 μM). The plates are left at room temperature for a further 2 hours before addition of 10 μl of the DiscoveRx PathHunter detection reagent (made up by adding 1 part Galacton Star, 5 parts Emerald II and 19 parts Cell Assay Buffer as per the manufacturer's instructions). After reagent addition, plates are covered and incubated at room temperature for 60 minutes. Luminescence signal is read using an Envision.
Measurement: IC50 = 22 nM (pChEMBL 7.659999847412109)
Descriptors: MW 568.44, ALogP 5.67, HBA 6, HBD 3, PSA 109.14, QED 0.28
Compound (SMILES): N#Cc1ccccc1CC(=O)NC1COc2ncccc21
InChIKey: BNGURZCIHJZDTH-UHFFFAOYSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: In Vitro Assay: Cell line generation was performed in general according to established molecular cloning protocols. Specifically, RNA was extracted from human whole blood using the Qiagen RNeasy kit (Qiagen, CH) according to the manufacturer's instructions. Subsequently cDNA was made (Superscript II, Invitrogen AG, CH) and the human P2X7 gene (genbank ref. BC011913) was amplified with the following primers: ATCGCGGCCGCTCAGTAAGGACTCTTGAAGCCACT and CGCCGCTAGCACCACCATGCCGGCCTGCTGCAGCTGCA. The amplified sequence was subsequently ligated into a pcDNA3.1 (+) NotI, NheI digested plasmid. Human embryonic kidney (HEK) cells (ATCC CRL-1573, Manassas, Va., USA) were transfected with the pcDNA3.1 (+). hP2X7 plasmid using lipofectamine 2000 (Invitrogen AG, CH) according to the manufacturer's instructions. Following a 24 h exposure to DNA, cells were trypsinized and re-seeded at low density in the presence of 250 ug Geneticin. Geneticin resistant cells were then selected during two consecutive rounds of cloning by serial limiting dilution with visual inspection.
Measurement: IC50 = 2975 nM (pChEMBL 5.53000020980835)
Descriptors: MW 279.3, ALogP 1.75, HBA 4, HBD 1, PSA 75.01, QED 0.93
Compound (SMILES): CC(C)C(=O)N1CC[C@H](CNC(=O)c2ccc(NC(=O)N3Cc4ccncc4C3)cc2)C1
InChIKey: HENBLOLUFWBSOW-QGZVFWFLSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Binding Assay : Probe 2: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-1287128.0. 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 = 5.67 nM (pChEMBL 8.25)
Descriptors: MW 435.53, ALogP 2.86, HBA 4, HBD 2, PSA 94.64, QED 0.76
Compound (SMILES): Nc1ncc(-c2cnn(CCN3CCOCC3)c2)cc1C(=O)Nc1ccncc1
InChIKey: KZXWFUQZOSWUCY-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase TBK1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UHD2
Assay: TBK1 : The kinase assay is performed as 384-well Flashplate assay assay (for e.g. Topcount measurement. 0.6 nM TANK binding kinase (TBK1), 800 nM biotinylated MELK-derived peptide (Biotin-Ah-Ah-AKPKGNKDYHLQTCCGSLAYRRR) SEQ ID NO: 2 and 10 uM ATP (spiked with 0.25 uCi 33P-ATP/well) are incubated in a total volume of 50 ul (10 mM MOPS, 10 mM Mg-acetat, 0.1 mM EGTA, 1 mM DTT, 0.02% Brij35, 0.1% BSA, pH 7.5) with or without test compound for 120 Min at 30° C. The reaction is stopped with 25 ul 200 mM EDTA. After 30 Min at room temperature the liquid is removed and each well washed thrice with 100 ul 0.9% sodium chloride solution.
Measurement: IC50 = 740 nM (pChEMBL 6.130000114440918)
Descriptors: MW 393.45, ALogP 1.51, HBA 8, HBD 2, PSA 111.19, QED 0.65
Compound (SMILES): CC(=O)Nc1nc2cc(-c3nn(Cc4cc5cccc(C)c5c(=O)n4-c4ccccc4C)c4ncnc(N)c34)ccc2o1
InChIKey: GQSFFKVELSYJJZ-UHFFFAOYSA-N
Target: PI3-kinase p110-alpha/p85-alpha (PROTEIN COMPLEX), Homo sapiens
UniProt: P42336
Assay: Inhibition of N-terminal His6-tagged recombinant full-length human p110alpha/untagged recombinant full length human p85alpha expressed in baculovirus infected Sf21 cells using phosphatidylinositol as substrate in presence of gamma-32P-ATP after 5 mins by thin layer chromatographic method
Measurement: IC50 < 10000 nM (pChEMBL 5)
Descriptors: MW 570.61, ALogP 5.14, HBA 10, HBD 2, PSA 146.75, QED 0.29
Compound (SMILES): O=C1C[C@@H](NC(=O)c2ccc3cc(Oc4ccc(C(F)(F)F)cn4)ccc3n2)CN1
InChIKey: TYWFTAYZYIPPMH-CYBMUJFWSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: FRET-Based Membrane Potential Assay: Two days prior to the experiment, frozen HEK293 cells stably expressing recombinant human Nav1.7 were quickly thawed and plated at 25,000 cells/well in growth medium [DMEM (Invitrogen #11965) with 10% HI FBS (Invitrogen #10082), 2 mM glutamine, 100 units/mL penicillin, 0.1 mg/mL streptomycin (PSG, Sigma #G1146), and 500 ug/mL Geneticin (Invitrogen #10131)] in black-walled, clear-bottom 384-well poly-D-lysine-coated assay plates (Greiner Bio-One, Frickenhausen, Germany) and incubated in a humidified 5% CO2 incubator at 37° C. On the day of the assay, medium was removed by aspiration, and cells were washed with assay buffer [HBSS (Invitrogen, Carlsbad, Calif.) containing 20 mM HEPES (Invitrogen, Carlsbad, Calif.)]. After washing, 30 uL assay buffer containing the fluorescent voltage-sensor probe CC2-DMPE (Invitrogen, Carlsbad, Calif.) at 20 uM and 0.01% pluronic F-127 (Invitrogen, Carlsbad, Calif.) was added to the cells. Cells were incubated for 40 minutes at room temperature in the dark. Following the incubation, the cells were washed and 30 uL assay buffer containing 2.5 uM DiSBAC2(3) substrate (Invitrogen, Carlsbad, Calif.) and 0.5 mM VABSC-1 (Invitrogen, Carlsbad, Calif.) was added to the cells. The cells were incubated for 90 minutes at room temperature in the dark. Fluorescence readings were made using a FLIPR TETRA (Molecular Devices, Sunnyvale Calif.) equipped with voltage-sensor probe optics. At the start of each experiment the optimal (EC80) concentration of depolarizing agent (veratridine) was determined by testing a dilution curve of assay buffer containing veratridine (Sigma-Aldrich, St. Louis, Mo.) and 1 mg/mL scorpion venom (SVqq, from Leiurus quinquestriatus; Sigma-Aldrich, St. Louis, Mo.). Compounds were dissolved in dimethyl sulfoxide, and 8-point, 1:3 dilution concentration-response curves were prepared in duplicate in dimethyl sulfoxide, followed by preparation of 0.8 uL/well daughter plates of the dilutions. Test compounds in the daughter plate were diluted to (Ë¿3x) solutions in assay buffer immediately before assaying. Using the FLIPR TETRA, 20 uL of the (3x) compound solutions were first added to the cells, then 20 uL of depolarizing solution (3xEC80 veratridine+SVqq) were added 3 minutes later to activate the channel. Changes in fluorescence were measured at wavelengths of 440-480 nm and 565-625 nm over the course of the experimental run. Membrane depolarization was expressed as a ratio of the maximum F440-480 nm/F565-625 nm reading above average baseline F440-480 nm/F565-625 nm reading. IC50 values were calculated from curve fits of the ratio data using a four-parameter logistic Hill equation (Accelrys Assay Explorer 3.3 Client, Accelrys, San Diego, Calif.) with percent inhibition plotted against compound concentration.
Measurement: IC50 = 1030 nM (pChEMBL 5.989999771118164)
Descriptors: MW 416.36, ALogP 3.06, HBA 5, HBD 2, PSA 93.21, QED 0.68
Compound (SMILES): O=C(OC1CCNC1)c1ccc2c(c1)-c1cc(Cl)c3cccnc3c1NS2(=O)=O
InChIKey: HHHISKQATYFMSQ-UHFFFAOYSA-N
Target: Ferroportin (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NP59
Assay: Antagonist activity at C-terminal halo-tag fused human ferroportin expressed in MDCK cells assessed as inhibition of hepcidin-induced ferroportin internalization after overnight incubation by Draq5 staining-based fluorescence assay
Measurement: IC50 = 25000 nM (pChEMBL 4.599999904632568)
Descriptors: MW 429.89, ALogP 3.19, HBA 6, HBD 2, PSA 97.39, QED 0.61
Compound (SMILES): COC(=O)Nc1nc2ccc(C(=O)c3ccccc3C(=O)NCc3ccsc3)cc2[nH]1
InChIKey: YIPLIXAWCFXUGF-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition of B-Raf (unknown origin) pre-incubated for 30 mins by luciferase-luciferin-coupled chemiluminescence assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 434.48, ALogP 3.96, HBA 6, HBD 3, PSA 113.18, QED 0.4
Compound (SMILES): CC1(C)CC(C2CCN(C(=O)c3ccc(N4CC(F)(F)C4)c(OCC4CC4)n3)CC2)C(=O)N1
InChIKey: IJYVQLCWSTUZBK-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×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 = 593.7 nM (pChEMBL 6.230000019073486)
Descriptors: MW 462.54, ALogP 3.09, HBA 5, HBD 1, PSA 74.77, QED 0.7
Compound (SMILES): COc1ccccc1OCCN1CCC(NS(=O)(=O)c2cncc3ccccc23)CC1
InChIKey: HBRGRVUSDCCMEP-UHFFFAOYSA-N
Target: Alpha-1A adrenergic receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35348
Assay: Intrinsic activity at alpha1A adrenergic receptor (unknown origin) assessed as inhibition of agonist induced effect preincubated for 30 mins followed by agonist addition measured after 5 hrs in presence of CCF4-AM by cell based beta lactamase reporter gene assay
Measurement: EC50 = 99.7 nM (pChEMBL 7)
Descriptors: MW 441.55, ALogP 3.07, HBA 6, HBD 1, PSA 80.76, QED 0.58
Compound (SMILES): COc1cc2oc(=O)cc(C)c2cc1-c1ccc(C#N)n1Cc1ccc(F)cc1
InChIKey: DYZJWTFYVLRGLB-UHFFFAOYSA-N
Target: Progesterone receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P06401
Assay: Antagonist activity at PR in human T47D cells assessed as inhibition of progesterone-induced alkaline phosphatase expression after 24 hrs by alkaline phopshatase assay
Measurement: IC50 = 570 nM (pChEMBL 6.239999771118164)
Descriptors: MW 388.4, ALogP 4.64, HBA 5, HBD 0, PSA 68.16, QED 0.48
Compound (SMILES): COC(=O)N[C@H](C(=O)N1CCC[C@H]1c1ncc(-c2ccc3c(c2)OCc2c-3c(=O)oc3cc(-c4cnc([C@@H]5CCCN5C(=O)[C@@H](NC(=O)OC)C(C)C)[nH]4)ccc23)[nH]1)C(C)C
InChIKey: ZAOOLKWRLJLQRV-RFOOMBAZSA-N
Target: Nonstructural protein 5A (SINGLE PROTEIN), Hepatitis C virus
UniProt: Q5L478
Assay: Inhibition of HCV genotype 1a NS5A Y93H mutant after 3 day incubation by RT-PCR-cell-based HCV replicon assay
Measurement: EC50 = 7 nM (pChEMBL 8.149999618530273)
Descriptors: MW 834.93, ALogP 6.22, HBA 11, HBD 4, PSA 214.08, QED 0.12