lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Cc1c(F)cccc1N1CCOc2cc(S(=O)(=O)Nc3nccs3)ccc21
InChIKey: NUASSICUZKIENU-UHFFFAOYSA-N
Target: Sodium channel protein type 5 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q14524
Assay: In Vitro Assay: HEK 293 cells stably transfected with either Nav 1.7 or with Nav 1.5 were recorded in population patch-clamp mode with the IonWorks® Quattro automated electrophysiology system in accordance with the manufacturer's specifications (Molecular Devices, LLC, Sunnyvale, Calif.). Sodium channel currents were measured in response to a train of depolarizations that induced successively greater inactivation. Cells were held at −110 mV for three seconds (Nav 1.7) or half a second (Nav 1.5) from a holding voltage of −15 mV, then put through a series of 26 pulses of 150 msec duration to −20 mV at a frequency of 5 Hz. Cells were then left unclamped for a period of 3 to 8 minutes while a single concentration of test compound was added. Cells were then reclamped and put through the same voltage protocol. Current at the end of the 26th pulse to −20 mV was subtracted from the peak current evoked by the 26th pulse to −20 mV to correct for leak current.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 405.48, ALogP 3.92, HBA 6, HBD 1, PSA 71.53, QED 0.71
Compound (SMILES): O=C1NCN([C@H]2CC[C@H](CNCC(F)(F)Br)CC2)c2c1cnc1[nH]ccc21
InChIKey: CDKRXWKJLOFKPT-HAQNSBGRSA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
Assay: Enzyme Assay: The inhibitory activity of the compounds of the present invention against JAK was measured.The respective enzymes (JAK1, JAK2, JAK3 and Tyk2) were purchased from Carna Biosciences, Inc.As a substrate of the enzymes (hereinafter referred to as substrate), LANCE Ultra ULight-JAK-1 (Tyr1023) Peptide (manufactured by PerkinElmer Inc.) was used.As an antibody for detecting phosphorylation of the substrate, LANCE Ultra Europium-anti-phospho tyrosine antibody (PT66) (manufactured by PerkinElmer Inc.) was used.The other reagents were purchased from the following.Adenosine triphosphate (ATP): Sigma-Aldrich4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES): DOJINDO LABORATORIESGlycol ether diamine tetraacetic acid (EGTA): DOJINDO LABORATORIES Magnesium chloride (MgCl2): Wako Pure Chemical Industries, Ltd.Dithiothreitol (DTT): Wako Pure Chemical Industries, Ltd.Tween 20: Sigma-AldrichEthylenediaminetetraacetic acid (EDTA): DOJINDO LABORATORIESThe compounds of the present invention, the enzymes (JAK1, JAK2, JAK3 and Tyk2), the substrate and ATP were used for assay as diluted with assay buffer.Â  As the assay buffer, one having the following composition was used.Â  HEPES (pH7.5): 50 mM EGTA: 1 mM MgCl2: 10 mM DTT: 2 mM Tween 20: 0.01% (wt/wt). The dilute concentration and the addition amount on a well plate as described hereinafter were adjusted so that the following final concentrations were achieved on the well plate.
Measurement: IC50 = 6.3 nM (pChEMBL 8.199999809265137)
Descriptors: MW 442.31, ALogP 3.21, HBA 4, HBD 3, PSA 73.05, QED 0.62
Compound (SMILES): C[C@@H]1CN([C@@H](C)CO)S(=O)(=O)c2ccc(C#Cc3cccc(F)c3)cc2O[C@H]1CN(C)C(=O)NC1CCCCC1
InChIKey: YAJYINBQFXCAPI-WENCSYSZSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Inhibition of human C-terminal His8-tagged IDH1-R132C mutant expressed in Escherichia coli Rosetta assessed as inhibition of ketoreductase activity by measuring NADPH consumption preincubated for 10 mins followed by addition of NADPH and alpha-KG as substrate by fluorescence method
Measurement: IC50 = 2500 nM (pChEMBL 5.599999904632568)
Descriptors: MW 571.71, ALogP 3.97, HBA 5, HBD 2, PSA 99.18, QED 0.53
Compound (SMILES): COc1cc(OC)cc(N(CCCNCC(F)F)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: YKAWIHSYTZKAKA-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 2754.23 nM (pChEMBL 5.559999942779541)
Descriptors: MW 482.54, ALogP 4.43, HBA 8, HBD 1, PSA 77.33, QED 0.32
Compound (SMILES): CC(=O)N1CCC(O)(C(=O)N2CC[C@](c3ccc(C(OCc4c(F)cccc4F)(C(F)(F)F)C(F)(F)F)cc3)(S(=O)(=O)c3ccc(F)cc3)C2)CC1
InChIKey: PIPPDPCSZQXYFH-HKBQPEDESA-N
Target: Nuclear receptor subfamily 1 group I member 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O75469
Assay: Transactivation of PXR in human HepG2 cells
Measurement: EC50 > 50000 nM (pChEMBL 4.301000118255615)
Descriptors: MW 766.68, ALogP 5.92, HBA 6, HBD 1, PSA 104.22, QED 0.23
Compound (SMILES): C[C@H](Nc1ncnc(N)c1C#Cc1cnccn1)c1nc2cccc(Cl)c2c(=O)n1-c1cccc(Cl)c1
InChIKey: RUCWAOMWOLHDQI-HNNXBMFYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Assay: PI3K isoforms were assayed under initial rate conditions in the presence of 25 mM Hepes (pH 7.4), and 2xKm ATP (100-300 uM), 10 uM PIP2, 5% glycerol, 5 mM MgCl2, 50 mM NaCl, 0.05% (v/v) Chaps, 1 mM dithiothreitol, 1% (v/v) DMSO at the following concentrations for each isoform: PI3K alpha, beta, delta, and gamma at 50 picomolar (pM) and PI3Kgamma at 2 nanomolar (nM). After an assay reaction time of 30 minutes at 25° C., reactions were terminated with a final concentration of 10 mM EDTA, 10 nM labeled-PIP3, and 35 nM Europium labeled GRP-1 detector protein before reading TR-FRET on an Envision plate reader (Ex: 340 nm; Em: 615/665 nm; 100 us delay and 500 us read window).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 529.39, ALogP 4.43, HBA 9, HBD 2, PSA 124.5, QED 0.33
Compound (SMILES): N#CC1=C(c2ccc3ccccc3c2)CC(c2ccc(OCCCC(F)(F)F)cc2)(C(F)(F)F)NC1=O
InChIKey: KQZJSFBORBDKNG-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: MGAT SPA Assay: MGAT2 enzyme was assayed using membranes isolated from Sf9 cells expressing the recombinant human MGAT2 cDNA with 2-monooleoylglycerol and [3H]-oleoyl-CoA as substrates as described by Seethala et al. [Anal. Biochem., 383(2):144-150 (Dec. 15, 2008)]. Briefly, the assays were conducted in 384-well plates in a total volume of 30 uL at 25° C. In each assay, 200 ng of recombinant human MGAT2 membrane was incubated with 10 uM of 2-monooleoylglycerol and 15 uM of [3H]-oleoyl-CoA in 100 mM potassium phosphate (pH 7.4) for 20 min with various concentrations of compounds delivered in DMSO. The assay was terminated by the addition of 20 ul of Stopping Solution (7.5 mg/ml Yttrium Oxide Polylysine beads, 3.3 mg/ml Fraction V BSA and 200 uM Mercuric chloride in 50 mM HEPES, pH 7.4). The signal was measured 1 h after quenching the reaction using LEADSEEKERSM for 5 minutes. To calculate the degree of inhibition, the zero level of enzyme activity (blank) was defined.
Measurement: IC50 = 684 nM (pChEMBL 6.170000076293945)
Descriptors: MW 518.46, ALogP 6.82, HBA 3, HBD 1, PSA 62.12, QED 0.29
Compound (SMILES): COc1cc2c(Nc3ccc(Br)c(C)c3)ncnc2cc1OCC1CN(C)CCO1
InChIKey: GFMZMGTWUAXGMZ-UHFFFAOYSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Inhibition of recombinant human N-terminal GST His6-tagged thrombin cleavage site-fused EGFR (H672 to A1210 residues) expressed in baculovirus using poly-EY as substrate after 210 mins by luciferase coupled chemiluminescence assay
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 473.37, ALogP 4.16, HBA 7, HBD 1, PSA 68.74, QED 0.58
Compound (SMILES): COc1c(Nc2ncc(Cl)c(Nc3ccccc3-n3cccn3)n2)ccc2c1CCCC(N1CCN(CCO)CC1)C2
InChIKey: RQTWMVOMNNMRAP-UHFFFAOYSA-N
Target: ALK tyrosine kinase receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UM73
Assay: Inhibition of NPM-ALK phosphorylation in human SUP-M2 cells after 2 to 3 hrs by ELISA
Measurement: IC50 = 20 nM (pChEMBL 7.699999809265137)
Descriptors: MW 589.14, ALogP 4.67, HBA 10, HBD 3, PSA 103.6, QED 0.24
Compound (SMILES): O=C(O)[C@H]1CC[C@](F)(C(=O)N2CC[C@](c3ccc(C(F)(C(F)(F)F)C(F)(F)F)cc3)(S(=O)(=O)c3ccc(F)cc3)C2)CC1
InChIKey: VXQLLPVKMJJWAK-GHEQTVGYSA-N
Target: Nuclear receptor subfamily 1 group I member 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O75469
Assay: Transactivation of PXR (unknown origin) expressed in human HepG2 cells
Measurement: EC50 = 10600 nM (pChEMBL 4.96999979019165)
Descriptors: MW 645.54, ALogP 6, HBA 4, HBD 1, PSA 91.75, QED 0.31
Compound (SMILES): CCCCCNC(=O)N(CCN1[C@@H]2CC[C@H]1C[C@@H](c1cccc(C(N)=O)c1)C2)CC1CCCCC1
InChIKey: GWDBIKQKJSMRNQ-LGKQTMLJSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: Displacement of [3H]-diprenorphine from human mu opioid receptor expressed in CHOK1 cell membranes
Measurement: Ki = 10 nM (pChEMBL 8)
Descriptors: MW 482.71, ALogP 5.28, HBA 3, HBD 2, PSA 78.67, QED 0.42
Compound (SMILES): IC[C@H]1CC[C@H](c2nnn3cnc4[nH]ccc4c23)CC1
InChIKey: OEBAWRRRIKOYQV-MGCOHNPYSA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
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 = 54 nM (pChEMBL 7.269999980926514)
Descriptors: MW 381.22, ALogP 3.31, HBA 4, HBD 1, PSA 58.87, QED 0.55
Compound (SMILES): COc1cccc(NC(=O)C2=C(c3ccc(C)cc3)CC(c3ccc(OCCCC(F)(F)F)cc3)(C(F)(F)F)NC2=O)c1
InChIKey: BYXAWQMDJIGRJC-UHFFFAOYSA-N
Target: Alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: Q10469
Assay: MGAT SPA Assay: MGAT2 enzyme was assayed using membranes isolated from Sf9 cells expressing the recombinant human MGAT2 cDNA with 2-monooleoylglycerol and [3H]-oleoyl-CoA as substrates as described by Seethala et al. [Anal. Biochem., 383(2):144-150 (Dec. 15, 2008)]. Briefly, the assays were conducted in 384-well plates in a total volume of 30 uL at 25° C. In each assay, 200 ng of recombinant human MGAT2 membrane was incubated with 10 uM of 2-monooleoylglycerol and 15 uM of [3H]-oleoyl-CoA in 100 mM potassium phosphate (pH 7.4) for 20 min with various concentrations of compounds delivered in DMSO. The assay was terminated by the addition of 20 ul of Stopping Solution (7.5 mg/ml Yttrium Oxide Polylysine beads, 3.3 mg/ml Fraction V BSA and 200 uM Mercuric chloride in 50 mM HEPES, pH 7.4). The signal was measured 1 h after quenching the reaction using LEADSEEKERSM for 5 minutes. To calculate the degree of inhibition, the zero level of enzyme activity (blank) was defined.
Measurement: IC50 = 275 nM (pChEMBL 6.559999942779541)
Descriptors: MW 606.56, ALogP 7.09, HBA 4, HBD 2, PSA 76.66, QED 0.15
Compound (SMILES): COc1cc2c(Nc3ccc(Br)c(Cl)c3Cl)ncnc2cc1OC[C@@H]1CO[C@H]2[C@@H]1OC[C@@H]2OC
InChIKey: QTWKCJAINLFWKV-ZMQOCNJZSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Inhibition of recombinant human N-terminal GST His6-tagged thrombin cleavage site-fused EGFR (H672 to A1210 residues) expressed in baculovirus using poly-EY as substrate after 210 mins by luciferase coupled chemiluminescence assay
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 571.26, ALogP 5.26, HBA 8, HBD 1, PSA 83.96, QED 0.39
Compound (SMILES): COc1cc(CS(C)(=O)=NC#N)cc(Nc2ncc(F)c(-c3ccc(F)cc3OC)n2)c1
InChIKey: BDRPEEVXRRASAS-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: P50750
Assay: CDK9/CycT1 : Recombinant full-length His-tagged human CDK9 and CycT1, expressed in insect cells and purified by Ni-NTA affinity chromatography, were purchase from Invitrogen (Cat. No PV4131). As substrate for the kinase reaction biotinylated peptide biotin-Ttds-YISPLKSPYKISEG (C-terminus in amid form) was used which can be purchased e.g. form the company JERINI peptide technologies (Berlin, Germany) For the assay 50 nl of a 100 fold concentrated solution of the test compound in DMSO was pipetted into a black low volume 384 well microtiter plate (Greiner Bio-One, Frickenhausen, Germany), 2 ul of a solution of CDK9/CycT1 in aqueous assay buffer [50 mM Tris/HCl pH 8.0, 10 mM MgCl2, 1.0 mM dithiothreitol, 0.1 mM sodium ortho-vanadate, 0.01% (v/v) Nonidet-P40 (Sigma)] were added and the mixture was incubated for 15 min at 22° C. to allow pre-binding of the test compounds to the enzyme before the start of the kinase reaction.
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 459.48, ALogP 4.26, HBA 8, HBD 1, PSA 109.49, QED 0.53
Compound (SMILES): COc1cc(N2CCN(CCS(C)(=O)=O)CC2)c(C)cc1Nc1nccc(-c2c(-c3cccc(C(=O)Nc4c(F)cccc4F)c3)nc3ccccn23)n1
InChIKey: LHSNLUJLSUOVPY-UHFFFAOYSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Inhibition of recombinant human full length EGFR expressed in baculovirus expression system assessed as decrease in transfer of gamma-phosphate from ATP onto tyrosine residue of biotinylated peptide substrate preincubated for 1 hr followed by substrate addition measured after 90 mins by HTRF method
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 752.85, ALogP 6.22, HBA 11, HBD 2, PSA 134.06, QED 0.16
Compound (SMILES): CCOc1cc(N2CCC(CCS(C)(=O)=O)CC2)c(C)cc1Nc1nccc(-c2c(-c3ccc(OC)c(C(=O)Nc4c(F)cccc4F)c3)nc3cc(F)ccn23)n1
InChIKey: HALNQUOIFUZUOJ-UHFFFAOYSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Inhibition of recombinant human full length EGFR expressed in baculovirus expression system assessed as decrease in transfer of gamma-phosphate from ATP onto tyrosine residue of biotinylated peptide substrate preincubated for 1 hr followed by substrate addition measured after 90 mins by HTRF method
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 813.9, ALogP 8.24, HBA 11, HBD 2, PSA 140.05, QED 0.12
Compound (SMILES): Cc1ccc(-c2ccc(Cl)c(F)c2)cc1C1=C(O)[C@]2(CC[C@H](C(F)(F)F)CC2)NC1=O
InChIKey: TZPOKOHORLSGEU-FVUUGDKNSA-N
Target: Acetyl-CoA carboxylase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O00763
Assay: Human ACC2 Enzyme Assay: For the assay, 50 nl of a 100-times concentrated solution of the test substance in DMSO were pipetted into a white low-volume 384-well microtitre plate (Greiner Bio-One, Frickenhausen, Germany), 2.5 μl of a solution of hACC2 in assay buffer [50 mM HEPES/NaOH pH 7.5, 2 mM MgCl2, 2 mM potassium citrate, 12 mM NaHCO3, 2 mM dithiothreitol (DTT), 0.005% (w/v) bovine serum albumin (BSA)] were added and the mixture was incubated for 15 min to allow prebinding of the substances to the enzyme prior to the enzyme reaction. The enzyme reaction was then started by addition of 2.5 μl of a solution of adenosine triphosphate (ATP, 100 μM=>final concentration in 5 μl of assay volume: 50 μM) and acetyl-CoA (20 μM=>final concentration in 5 μl assay volume: 10 μM) in assay buffer, and the resulting mixture was incubated at 22 °C. for the reaction time of 45 min. The concentration of the hACC2 was adjusted to the respective activity of the enzyme and set such that the assay was carried out in the linear range. Typical concentrations were in the range of 2 ng/μl. The reaction was stopped by addition of 2.5 μl of the "ADP-GLO reagent" (1:1.5-times diluted), and the resulting mixture was incubated at 22 °C. for 1 h to convert the unreacted ATP completely into cAMP. 2.5 μl of the "kinase detection reagent" were then added (1.2-times more concentrated than recommended by the manufacturer), the resulting mixture was incubated at 22 °C. for 1 h and the luminescence was then measured using a suitable measuring instrument (Viewlux or Topcount from Perkin-Elmer or Pherastar from BMG Labtechnologies).
Measurement: IC50 = 580 nM (pChEMBL 6.239999771118164)
Descriptors: MW 453.86, ALogP 6.34, HBA 2, HBD 2, PSA 49.33, QED 0.52
Compound (SMILES): COc1cc2c(Nc3ccc(Cl)c(Cl)c3)ncnc2cc1OCC1CN(Cc2ccccc2)CCO1
InChIKey: QUDDVDXFQIWBKM-UHFFFAOYSA-N
Target: Ephrin type-A receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29317
Assay: AlphaScreen Assay: In AlphaScreen, when the donor and acceptor beads are close in proximity, a series of photochemical events will give rise to a fluorescent signal upon light activation. Here we use biotinylated poly-(Glu, Tyr) 4:1 as the kinase substrate, streptavidin-coated donor beads and anti-phosphortyrosine antibody PY100-coated acceptor beads. Upon phosphorylation, the peptide substrate can bind to both donor and acceptor beads, thus gives rise to fluorescence. Compounds, ATP, biotinylated poly-(Glu, Tyr) and kinases are mixed in a volume of 20 μL for 1 hr at ambient temperature using Greiner 384-well white clear bottom medium binding plates. Then 10 uL solution containing 15-30 mg/mL AlphaScreen beads, 75 mM Hepes, pH 7.4, 300 mM NaCl, 120 mM EDTA, 0.3% BSA and 0.03% Tween-20 is added to each well. After 2-16 hr incubation of the beads, plates are read in a Perkin Elmer AlphaQuest reader (available from PerkinElmer Life and Analytical Sciences, Inc., of Boston Mass.).
Measurement: IC50 = 10500 nM (pChEMBL 4.980000019073486)
Descriptors: MW 525.44, ALogP 5.97, HBA 7, HBD 1, PSA 68.74, QED 0.3
Compound (SMILES): O=S(=O)(Nc1ccncn1)c1ccc2c(c1)OCCN2c1ccc(C(F)(F)F)cc1C1=CCNCC1
InChIKey: FQJDSGMAKKSCMD-UHFFFAOYSA-N
Target: Sodium channel protein type 5 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q14524
Assay: In Vitro Assay: HEK 293 cells stably transfected with either Nav 1.7 or with Nav 1.5 were recorded in population patch-clamp mode with the IonWorks® Quattro automated electrophysiology system in accordance with the manufacturer's specifications (Molecular Devices, LLC, Sunnyvale, Calif.). Sodium channel currents were measured in response to a train of depolarizations that induced successively greater inactivation. Cells were held at −110 mV for three seconds (Nav 1.7) or half a second (Nav 1.5) from a holding voltage of −15 mV, then put through a series of 26 pulses of 150 msec duration to −20 mV at a frequency of 5 Hz. Cells were then left unclamped for a period of 3 to 8 minutes while a single concentration of test compound was added. Cells were then reclamped and put through the same voltage protocol. Current at the end of the 26th pulse to −20 mV was subtracted from the peak current evoked by the 26th pulse to −20 mV to correct for leak current.
Measurement: IC50 = 1200 nM (pChEMBL 5.920000076293945)
Descriptors: MW 517.53, ALogP 4.2, HBA 7, HBD 2, PSA 96.45, QED 0.52
Compound (SMILES): CN(C)C1(c2ccccc2)CCC(c2ccccc2)(N(C)Cc2ccccc2)CC1
InChIKey: JVSMGZQUZMVUJJ-UHFFFAOYSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: Binding Assay: The affinity to the human μ-opiate receptor was determined in a homogeneous preparation in microtiter plates. For this, dilution series of the respective compound to be tested were incubated for 90 minutes at room temperature with a receptor membrane preparation (15-40 mg of protein per 250 μl of incubation batch) of CHO-K 1 cells, which express the human μ-opiate receptor (RB-HOM receptor membrane preparation of NEN, Zaventem, Belgium), in the presence of 1 nmol/l of the radioactive ligand [3H'-naloxone (NET719, NEN, Zaventem, Belgium) and of 1 mg WGA-SPA beads (wheat germ agglutinin SPA beads from Amersham/Pharmacia, Freiburg, Germany) in a total volume of 250 μl. 50 mmol/l of tris-HCl supplemented by 0.05% by wt. of sodium azide and 0.06% by wt. of bovine serum albumin was used as incubation buffer. 25 μmol/l of naloxone were additionally added to determine the non-specific bond. After the ninety-minute incubation time had ended, the microtiter plates were centrifuged for 20 minutes at 1000 g and the radioactivity measured in a β-counter (Microbeta-Trilux, PerkinElmer Wallac, Freiburg, Germany).
Measurement: Ki = 120 nM (pChEMBL 6.920000076293945)
Descriptors: MW 398.59, ALogP 6.05, HBA 2, HBD 0, PSA 6.48, QED 0.5
Compound (SMILES): OCCN(C[C@H]1CC[C@H](c2nnn3cnc4[nH]ccc4c23)CC1)C1CCCCC1
InChIKey: PYELFLISDANEIM-QAQDUYKDSA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
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 > 1000 nM (pChEMBL 6)
Descriptors: MW 396.54, ALogP 3.51, HBA 6, HBD 2, PSA 82.34, QED 0.67
Compound (SMILES): Cc1cccc2cc(Cn3nc(C(C)(C)O)c4c(N)ncnc43)n(-c3ccccc3)c(=O)c12
InChIKey: LLOJBGUZWFEGFN-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 440.51, ALogP 3.3, HBA 8, HBD 2, PSA 111.85, QED 0.44
Compound (SMILES): Cc1cccc2cc([C@H](C)Nc3ncnc4[nH]nc(Cl)c34)n(C3CC3)c(=O)c12
InChIKey: GWRXJSIAEKYOIY-NSHDSACASA-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 394.87, ALogP 4.14, HBA 6, HBD 2, PSA 88.49, QED 0.54
Compound (SMILES): Cc1cccc2cc(Cn3nc(/C=C4/CCNC4=O)c4c(N)ncnc43)n(-c3ccccc3F)c(=O)c12
InChIKey: AWMKMMUIWQVKMO-ATVHPVEESA-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 495.52, ALogP 3.11, HBA 8, HBD 2, PSA 120.72, QED 0.37
Compound (SMILES): COC(=O)Nc1ccc(-c2[nH]c([C@@H]3C[C@H](C4CCN(S(C)(=O)=O)CC4)CN3C(=O)[C@H]3CC[C@H](CN)CC3)nc2Cl)cc1
InChIKey: SFHOTXRQRGQBBC-CKLTXHEASA-N
Target: Coagulation factor X (SINGLE PROTEIN), Homo sapiens
UniProt: P00742
Assay: In Vitro Assay: Human Factor Xa (American Diagnostica Inc.) and human thrombin (Sigma) activities were measured at the enzyme concentrations of 0.18 U/mL and 0.12 U/mL, respectively in the same buffer containing 150 mM NaCl, 2 mg/mL PEG6000, 50 mM Tris-HCl (pH7.4), except that the reactions were started with 300 μM S-2222 (phenyl-Ile-Glu-Gly-Arg-pNA, Chromogenix) and 300 μM S-2366, respectively.
Measurement: IC50 > 100000 nM (pChEMBL 4)
Descriptors: MW 621.2, ALogP 4.23, HBA 7, HBD 3, PSA 150.72, QED 0.42
Compound (SMILES): O=S(=O)(Nc1ccncn1)c1ccc2c(c1)OCCN2c1ncc(C(F)(F)F)cc1Cl
InChIKey: QZKFAPZUVJIKCJ-UHFFFAOYSA-N
Target: Sodium channel protein type 5 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q14524
Assay: In Vitro Assay: HEK 293 cells stably transfected with either Nav 1.7 or with Nav 1.5 were recorded in population patch-clamp mode with the IonWorks® Quattro automated electrophysiology system in accordance with the manufacturer's specifications (Molecular Devices, LLC, Sunnyvale, Calif.). Sodium channel currents were measured in response to a train of depolarizations that induced successively greater inactivation. Cells were held at −110 mV for three seconds (Nav 1.7) or half a second (Nav 1.5) from a holding voltage of −15 mV, then put through a series of 26 pulses of 150 msec duration to −20 mV at a frequency of 5 Hz. Cells were then left unclamped for a period of 3 to 8 minutes while a single concentration of test compound was added. Cells were then reclamped and put through the same voltage protocol. Current at the end of the 26th pulse to −20 mV was subtracted from the peak current evoked by the 26th pulse to −20 mV to correct for leak current.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 471.85, ALogP 3.88, HBA 7, HBD 1, PSA 97.31, QED 0.62
Compound (SMILES): COc1ccc(N2CCOc3cc(S(=O)(=O)Nc4nccs4)ccc32)cc1C#N
InChIKey: MKPZIIPTMFMOCV-UHFFFAOYSA-N
Target: Sodium channel protein type 5 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q14524
Assay: In Vitro Assay: HEK 293 cells stably transfected with either Nav 1.7 or with Nav 1.5 were recorded in population patch-clamp mode with the IonWorks® Quattro automated electrophysiology system in accordance with the manufacturer's specifications (Molecular Devices, LLC, Sunnyvale, Calif.). Sodium channel currents were measured in response to a train of depolarizations that induced successively greater inactivation. Cells were held at −110 mV for three seconds (Nav 1.7) or half a second (Nav 1.5) from a holding voltage of −15 mV, then put through a series of 26 pulses of 150 msec duration to −20 mV at a frequency of 5 Hz. Cells were then left unclamped for a period of 3 to 8 minutes while a single concentration of test compound was added. Cells were then reclamped and put through the same voltage protocol. Current at the end of the 26th pulse to −20 mV was subtracted from the peak current evoked by the 26th pulse to −20 mV to correct for leak current.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 428.5, ALogP 3.35, HBA 8, HBD 1, PSA 104.55, QED 0.66
Compound (SMILES): O=C(NO)c1cnc(NC2(c3ccccc3)CCN(C(=O)Nc3cccc(F)c3)CC2)nc1
InChIKey: AQAFOMARFMGXDU-UHFFFAOYSA-N
Target: Histone deacetylase 3 (SINGLE PROTEIN), Homo sapiens
UniProt: O15379
Assay: HDAC Enzyme Assay: The compounds were diluted in assay buffer (50 mM HEPES, pH 7.4, 100 mM KCl, 0.001% Tween-20, 0.05% BSA, 20 uM TCEP) to 6 fold their final concentration. The HDAC enzymes (purchased from BPS Biosciences) were diluted to 1.5 fold their final concentration in assay buffer. The tripeptide substrate and trypsin at 0.05 uM final concentration were diluted in assay buffer at 6 fold their final concentration. The final enzyme concentrations used in these assays were 3.3 ng/ml (HDAC1), 0.2 ng/ml (HDAC2), 0.08 ng/ml (HDAC3) and 2 ng/ml (HDAC6). The final substrate concentrations used were 16 uM (HDAC1), 10 uM (HDAC2), 17 uM (HDAC3) and 14 uM (HDAC6). Five ul of compounds and 20 ul of enzyme were added to wells of a black, opaque 384 well plate in duplicate. Enzyme and compound were incubated together at room temperature for 10 minutes. Five u1 of substrate was added to each well, the plate was shaken for 60 seconds and placed into a Victor 2 microtiter plate reader. The development of fluorescence was monitored for 60 min and the linear rate of the reaction was calculated.
Measurement: IC50 = 1081 nM (pChEMBL 5.96999979019165)
Descriptors: MW 450.47, ALogP 3.37, HBA 6, HBD 4, PSA 119.48, QED 0.35
Compound (SMILES): Nc1cc(F)ccc1NC(=O)CCCCCNS(=O)(=O)c1ccc(F)cc1
InChIKey: VYACAXJVXSLSDO-UHFFFAOYSA-N
Target: Histone deacetylase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13547
Assay: HDAC enzyme inhibitionAssay: HDAC enzyme inhibition assays were performed using purified HDACs 1-10 essentially as described in Beckers et al., 2007, Int. J. Cancer., 121:1138-48 and Perez-Balado et al., 2007, J. Med. Chem., 50:2497-2505. Inhibition assays using nuclear extract were performed essentially as described in Herman et al., 2006, Nat. Chem. Biol., 2:551-558. Briefly, the purified HDACs or nuclear extract were incubated with an acetylated substrate in the absence of the compound to be assayed and with increasing concentrations of the compound. The rate of substrate deacetylation was measured under each condition, and half-maximal inhibitory concentration with regard to each HDAC was determined by standard means.
Measurement: IC50 = 30000 nM (pChEMBL 4.519999980926514)
Descriptors: MW 397.45, ALogP 3.02, HBA 4, HBD 3, PSA 101.29, QED 0.45
Compound (SMILES): COc1cc(F)ccc1-c1nc(Nc2cc(F)cc(CS(=N)(=O)C3CC3)c2)ncc1F
InChIKey: XDPKOVYWCYQSBM-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: P50750
Assay: CDK9/CycT1 Kinase Assay: Recombinant full-length His-tagged human CDK9 and CycT1, expressed in insect cells and purified by Ni-NTA affinity chromatography, were purchased from Invitrogen (Cat. No PV4131). As substrate for the kinase reaction biotinylated peptide biotin-Ttds-YISPLKSPYKISEG (C-terminus in amid form) was used which can be purchased e.g. form the company JERINI Peptide Technologies (Berlin, Germany) For the assay 50 nl of a 100 fold concentrated solution of the test compound in DMSO was pipetted into a black low volume 384 well microtiter plate (Greiner Bio-One, Frickenhausen, Germany), 2 ul of a solution of CDK9/CycT1 in aqueous assay buffer [50 mM Tris/HCl pH 8.0, 10 mM MgCl2, 1.0 mM dithiothreitol, 0.1 mM sodium ortho-vanadate, 0.01% (v/v) Nonidet-P40 (Sigma)] were added and the mixture was incubated for 15 min at 22 C. to allow pre-binding of the test compounds to the enzyme before the start of the kinase reaction.
Measurement: IC50 = 7 nM (pChEMBL 8.149999618530273)
Descriptors: MW 448.47, ALogP 5.02, HBA 6, HBD 2, PSA 87.96, QED 0.53
Compound (SMILES): COc1cc(OC)cc(N(CCCNC(C)C)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1.Cl
InChIKey: NWRFMCQOAWFURR-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 1288.25 nM (pChEMBL 5.889999866485596)
Descriptors: MW 460.58, ALogP 4.57, HBA 8, HBD 1, PSA 77.33, QED 0.35
Compound (SMILES): COc1c(Nc2ncc(Cl)c(Nc3ccccc3N(C)S(C)(=O)=O)n2)ccc2c1CCCC(N1CCN(CCO)CC1)C2
InChIKey: QUZLPRLQAANSNR-UHFFFAOYSA-N
Target: ALK tyrosine kinase receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UM73
Assay: Inhibition of NPM-ALK phosphorylation in human SUP-M2 cells after 2 to 3 hrs by ELISA
Measurement: IC50 = 30 nM (pChEMBL 7.519999980926514)
Descriptors: MW 630.22, ALogP 3.88, HBA 10, HBD 3, PSA 123.16, QED 0.28
Compound (SMILES): COc1ccc(-c2cn3c4nc(ncc24)NCCCCCCCC(O)CCCC3)cc1
InChIKey: XXEJLOWSXIMQHG-UHFFFAOYSA-N
Target: Tyrosine-protein kinase receptor TYRO3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q06418
Assay: Inhibition of Tyro3 (unknown origin) by microfluidic capillary electrophoresis assay
Measurement: IC50 = 940 nM (pChEMBL 6.03000020980835)
Descriptors: MW 422.57, ALogP 5.4, HBA 6, HBD 2, PSA 72.2, QED 0.58
Compound (SMILES): CC(C)COc1ncnc2[nH]cc(/C=C/C(=O)NC3CC3)c12
InChIKey: NTUWMPKGQOZJLQ-ZZXKWVIFSA-N
Target: TGF-beta-activated kinase 1 and MAP3K7-binding protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q15750
Assay: Biological Assay: TAK1-TAB1 Binding Inhibitory Potency: The ability of candidate compounds to interact with TAK1-TAB1 is quantitated by a competitive binding assay using the LanthaScreen technology developed by Life Technologies. This assay is based on the binding of a proprietary, Alexa Fluor 647-labeled, ATP-competitive kinase inhibitor (kinase tracer-236) to the TAK1-TAB1 construct in presence of a europium-conjugated antibody, resulting in a FRET (fluorescence resonance energy transfer) signal. Displacement of the kinase tracer by compound results in a lower emission ratio upon excitation of the europium chelate. Candidate compounds are designed as potential irreversible inhibitors of TAK1-TAB1, capable of ligating to an active site cysteine residue. The time dependent nature of irreversible inhibition is investigated by performing the binding assay with and without a pre-incubation of compound and TAK1-TAB1. An increase in potency in the pre-incubated assay suggests the candidate compound could b of 10 nM TAK1-TAB1, 2 nM Eu-anti-his antibody, and 100 nM kinase tracer-236 using a 384-well plate format. Background signal is defined in the absence of TAK1-TAB1 and uninhibited signal is defined in the presence of vehicle (2% DMSO) alone. Compounds were evaluated in an 11 point dose-response ranging from 20 uM to 0.34 nM. The binding assays are performed under two conditions to evaluate time dependence of inhibition. For the pre-incubation assay, TAK1-TAB1 and Eu-anti-his antibody are preincubated with compound or vehicle for two hours prior to the addition of kinase tracer. The non-preincubated assay is run in which TAK1-TAB1 and Eu-anti-his antibody are added to a mixture of compound and kinase tracer. IC50 values of compounds are determined using a 4 parameter logistical fit of emission ratio as a function of the concentration of compound.
Measurement: IC50 = 180 nM (pChEMBL 6.75)
Descriptors: MW 300.36, ALogP 2.28, HBA 4, HBD 2, PSA 79.9, QED 0.8
Compound (SMILES): Cc1cncc(-c2cc3c(cn2)cnn3-c2cccc(N3CC(O)C(O)C3)n2)n1
InChIKey: VDJIRBSRWJZSCK-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q86V86
Assay: Kinase Binding Assay: PIM-1, -2, and -3 enzymes were generated as fusion proteins expressed in bacteria and purified by IMAC column chromatography (Sun, X., Chiu, J. F., and He, Q. Y. (2005) Expert Rev. Proteomics, 2:649-657). A fluorescent-labeled Pim-specific peptide substrate, was custom synthesized by American Peptide Company (Sunnyvale, Calif.). Reaction Buffer contained 10 mM HEPES, pH 7.2, 10 mM MgCl2, 0.01% Tween 20, 2 mM DTT. Termination Buffer contained 190 mM HEPES, pH 7.2, 0.015% Brij-35, 0.2% Coating Reagent 3 (Caliper Life Sciences, Hopkinton, Mass.), 20 mM EDTA. Separation Buffer contained 100 mM HEPES, pH 7.2, 0.015% Brij-35, 0.1% Coating Reagent 3, 1:200 Coating Reagent 8 (Caliper Life Sciences, Hopkinton, Mass.), 10 mM EDTA and 5% DMSO.PIM reactions were carried out in a final volume of 10 uL per well in a 384-well plate. A standard enzymatic reaction, initiated by the addition of 5 uL 2Ã¿Â¿ATP and test compound to 5 uL of 2x enzyme and FAM-peptide, contained 20 uM PIM1, 50 uM PIM2, or 55 uM PIM3, 1 uM FAM-peptide, and 10 uM ATP, in Reaction Buffer. After 90 minutes of incubation at room temperature, the phosphorylation reaction was stopped by the addition of 10 uL Termination Buffer. The product and substrate in each independent reaction were separated on a 12-sipper microfluidic chip (Caliper Life Sciences, Hopkinton, Mass.) run on a Caliper LC3000 (Caliper Life Sciences, Hopkinton, Mass.). The separation of product and substrate was optimized by choosing voltages and pressure using Caliper's Optimizer software (Hopkinton, Mass.). The separation conditions used a downstream voltage of -500V, an upstream voltage of -2150V, and a screening pressure of -1.2 psi. The product and substrate fluorophore were excited at 488 nm and detected at 530 nm. Substrate conversion was calculated from the electropherogram using HTS Well Analyzer software (Caliper Life Sciences, Hopkinton, Mass.). Ki values for the test compound were calculated.
Measurement: Ki = 0.83 nM (pChEMBL 9.079999923706055)
Descriptors: MW 389.42, ALogP 1.12, HBA 9, HBD 2, PSA 113.08, QED 0.54
Compound (SMILES): Cc1ccc(Cl)cc1N1CCOc2cc(S(=O)(=O)Nc3nccs3)ccc21
InChIKey: BXNORRGHXDHNQQ-UHFFFAOYSA-N
Target: Sodium channel protein type 5 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q14524
Assay: In Vitro Assay: HEK 293 cells stably transfected with either Nav 1.7 or with Nav 1.5 were recorded in population patch-clamp mode with the IonWorks® Quattro automated electrophysiology system in accordance with the manufacturer's specifications (Molecular Devices, LLC, Sunnyvale, Calif.). Sodium channel currents were measured in response to a train of depolarizations that induced successively greater inactivation. Cells were held at −110 mV for three seconds (Nav 1.7) or half a second (Nav 1.5) from a holding voltage of −15 mV, then put through a series of 26 pulses of 150 msec duration to −20 mV at a frequency of 5 Hz. Cells were then left unclamped for a period of 3 to 8 minutes while a single concentration of test compound was added. Cells were then reclamped and put through the same voltage protocol. Current at the end of the 26th pulse to −20 mV was subtracted from the peak current evoked by the 26th pulse to −20 mV to correct for leak current.
Measurement: IC50 = 5600 nM (pChEMBL 5.25)
Descriptors: MW 421.93, ALogP 4.44, HBA 6, HBD 1, PSA 71.53, QED 0.67
Compound (SMILES): Cc1cncc(-c2cc3c(cn2)cnn3-c2ccc(C(F)(F)F)c(N3CCNCC(O)C3)n2)n1
InChIKey: RVQPBMWIXPQVNW-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q86V86
Assay: Kinase Binding Assay: PIM-1, -2, and -3 enzymes were generated as fusion proteins expressed in bacteria and purified by IMAC column chromatography (Sun, X., Chiu, J. F., and He, Q. Y. (2005) Expert Rev. Proteomics, 2:649-657). A fluorescent-labeled Pim-specific peptide substrate, was custom synthesized by American Peptide Company (Sunnyvale, Calif.). Reaction Buffer contained 10 mM HEPES, pH 7.2, 10 mM MgCl2, 0.01% Tween 20, 2 mM DTT. Termination Buffer contained 190 mM HEPES, pH 7.2, 0.015% Brij-35, 0.2% Coating Reagent 3 (Caliper Life Sciences, Hopkinton, Mass.), 20 mM EDTA. Separation Buffer contained 100 mM HEPES, pH 7.2, 0.015% Brij-35, 0.1% Coating Reagent 3, 1:200 Coating Reagent 8 (Caliper Life Sciences, Hopkinton, Mass.), 10 mM EDTA and 5% DMSO.PIM reactions were carried out in a final volume of 10 uL per well in a 384-well plate. A standard enzymatic reaction, initiated by the addition of 5 uL 2Ã¿Â¿ATP and test compound to 5 uL of 2x enzyme and FAM-peptide, contained 20 uM PIM1, 50 uM PIM2, or 55 uM PIM3, 1 uM FAM-peptide, and 10 uM ATP, in Reaction Buffer. After 90 minutes of incubation at room temperature, the phosphorylation reaction was stopped by the addition of 10 uL Termination Buffer. The product and substrate in each independent reaction were separated on a 12-sipper microfluidic chip (Caliper Life Sciences, Hopkinton, Mass.) run on a Caliper LC3000 (Caliper Life Sciences, Hopkinton, Mass.). The separation of product and substrate was optimized by choosing voltages and pressure using Caliper's Optimizer software (Hopkinton, Mass.). The separation conditions used a downstream voltage of -500V, an upstream voltage of -2150V, and a screening pressure of -1.2 psi. The product and substrate fluorophore were excited at 488 nm and detected at 530 nm. Substrate conversion was calculated from the electropherogram using HTS Well Analyzer software (Caliper Life Sciences, Hopkinton, Mass.). Ki values for the test compound were calculated.
Measurement: Ki = 0.265 nM (pChEMBL 9.579999923706055)
Descriptors: MW 470.46, ALogP 2.37, HBA 9, HBD 2, PSA 104.88, QED 0.47
Compound (SMILES): CC(C)[C@H](NC(=O)C(=O)Nc1cccc2ccccc12)C(=O)NC(CC(=O)O)C(=O)CF
InChIKey: DCOFJYKNJPKGOK-CVMIBEPCSA-N
Target: Caspase-6 (SINGLE PROTEIN), Homo sapiens
UniProt: P55212
Assay: Inhibition of recombinant MCH2 (unknown origin) pre-incubated for 60 mins with Acetyl-Tyr-Val-Ala-Asp-amino-4-methylcoumarin substrate followed by enzyme addition by fluorescence based assay
Measurement: Ki = 17 nM (pChEMBL 7.769999980926514)
Descriptors: MW 445.45, ALogP 1.42, HBA 5, HBD 4, PSA 141.67, QED 0.43
Compound (SMILES): N#Cc1cccc(S(=O)(=O)Nc2ccc(-c3nc4cc(NS(=O)(=O)c5cccc(C#N)c5)ccc4o3)cc2)c1
InChIKey: GKIROPJDTJPOEB-UHFFFAOYSA-N
Target: HSP60/HSP10 (PROTEIN COMPLEX), Homo sapiens
UniProt: P61604
Assay: Inhibition of human mitochondrial HSP60/HSP10 expressed in Escherichia coli DH5alpha/BL21 (DE3) assessed as inhibition of denatured MDH refolding preincubated for 10 mins followed by ATP addition measured after 40 to 60 mins by fluorescence based assay
Measurement: IC50 = 91000 nM (pChEMBL 4.039999961853027)
Descriptors: MW 555.6, ALogP 4.84, HBA 8, HBD 2, PSA 165.95, QED 0.29
Compound (SMILES): COc1cc(OC)cc(N(CCN2CCNC(=O)C2)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: VIADTQYJXCPMEC-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 4570.88 nM (pChEMBL 5.340000152587891)
Descriptors: MW 487.56, ALogP 2.62, HBA 9, HBD 1, PSA 97.64, QED 0.41
Compound (SMILES): NC(Cc1cc(-c2cccc(C(=O)O)c2)cc([N+](=O)[O-])c1)C(=O)O
InChIKey: SXFXSXBKFUFSCC-UHFFFAOYSA-N
Target: Glutamate receptor ionotropic, kainate 3 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P42264
Assay: Displacement of [3H]-kainate from rat recombinant GluK3 receptor expressed in baculovirus infected insect Sf9 cell membranes after 60 mins
Measurement: Ki > 100000 nM (pChEMBL 4)
Descriptors: MW 330.3, ALogP 1.91, HBA 5, HBD 3, PSA 143.76, QED 0.54
Compound (SMILES): N#CC1CCCN1C(=O)CNC(=O)c1cccc2cnccc12
InChIKey: DSIXHJMXKMIOFX-UHFFFAOYSA-N
Target: Dipeptidyl peptidase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: Q86TI2
Assay: Enzymatic Assay: Enzyme activities were determined kinetically in a final volume of 200 μl for 10 minutes at 37° C. by measuring the initial velocities of pNA release (405 nm) from the substrate using a Spectramax plus microtiterplate reader (Molecular devices). One unit of enzyme activity was defined as the amount of enzyme that catalyzes the release of 1 μmol pNA from the substrate per minute under assay conditions. The chromogenic substrate Gly-Pro-p-nitroanilide (100 μmol/l) was used at pH 8.3 for DPP IV, Lys-Ala-p-nitroanilide (1 mmol/l) at pH 5.5 for DPP II, Ala-Pro-p-nitroanilide (300 μmol/l) at pH 7.4 for DPP9 and Ala-Pro-p-nitroanilide (2 mmol/l) at pH 7.4 for FAP activity measurement. To evaluate the endopeptidase activity of FAP and the influence of inhibitors thereon, Z-Gly-Pro-AMC and Z-Gly-Pro-p-nitroanilide were used at a final concentration of 300 and 100 μmol/l, respectively. The substrate concentrations were chosen around the Km value obtained under the assay conditions used. Buffer compositions for the DPP assays were reported before in the purification articles−vide supra. The FAP assay buffer consisted of 50 mM Tris pH7.4 containing 100 mmol/l NaCl and 0.1 mg/ml bovine serum albumin. The PREP activity was measured as described by Brandt et al. using the chromogenic substrate Z-Gly-Pro-p-nitroanilide (0.25 mmol/l) at pH 7.5 in the presence of 10 mmol/l DTT. Test compounds were dissolved and diluted in DMSO (final concentration DMSO during assay 5% v/v) except for FAP where dilution of the inhibitor was done in water. Inhibitors are pre-incubated with the enzyme for 15 min at 37° C. before starting the assay by the addition of substrate. The concentration of enzyme and of inhibitor during the preincubation is double of the final concentration during activity measurement.
Measurement: IC50 > 100000 nM (pChEMBL 4)
Descriptors: MW 308.34, ALogP 1.48, HBA 4, HBD 1, PSA 86.09, QED 0.93
Compound (SMILES): COc1cc(N2CCC(N3CCCCC3)CC2)c(C)cc1Nc1nccc(-c2c(-c3cccc(C(=O)Nc4c(F)cccc4F)c3)nc3ccccn23)n1
InChIKey: GPOXREFACYVEDP-UHFFFAOYSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: Inhibition of recombinant human full length EGFR expressed in baculovirus expression system assessed as decrease in transfer of gamma-phosphate from ATP onto tyrosine residue of biotinylated peptide substrate preincubated for 1 hr followed by substrate addition measured after 90 mins by HTRF method
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 728.85, ALogP 8.5, HBA 9, HBD 2, PSA 99.92, QED 0.15
Compound (SMILES): OCC#Cc1cncc(-c2cc3c(cn2)cnn3-c2cccc(N3CCNCC3)n2)n1
InChIKey: AZUDXPDVDQXJJH-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q86V86
Assay: Kinase Binding Assay: PIM-1, -2, and -3 enzymes were generated as fusion proteins expressed in bacteria and purified by IMAC column chromatography (Sun, X., Chiu, J. F., and He, Q. Y. (2005) Expert Rev. Proteomics, 2:649-657). A fluorescent-labeled Pim-specific peptide substrate, was custom synthesized by American Peptide Company (Sunnyvale, Calif.). Reaction Buffer contained 10 mM HEPES, pH 7.2, 10 mM MgCl2, 0.01% Tween 20, 2 mM DTT. Termination Buffer contained 190 mM HEPES, pH 7.2, 0.015% Brij-35, 0.2% Coating Reagent 3 (Caliper Life Sciences, Hopkinton, Mass.), 20 mM EDTA. Separation Buffer contained 100 mM HEPES, pH 7.2, 0.015% Brij-35, 0.1% Coating Reagent 3, 1:200 Coating Reagent 8 (Caliper Life Sciences, Hopkinton, Mass.), 10 mM EDTA and 5% DMSO.PIM reactions were carried out in a final volume of 10 uL per well in a 384-well plate. A standard enzymatic reaction, initiated by the addition of 5 uL 2Ã¿Â¿ATP and test compound to 5 uL of 2x enzyme and FAM-peptide, contained 20 uM PIM1, 50 uM PIM2, or 55 uM PIM3, 1 uM FAM-peptide, and 10 uM ATP, in Reaction Buffer. After 90 minutes of incubation at room temperature, the phosphorylation reaction was stopped by the addition of 10 uL Termination Buffer. The product and substrate in each independent reaction were separated on a 12-sipper microfluidic chip (Caliper Life Sciences, Hopkinton, Mass.) run on a Caliper LC3000 (Caliper Life Sciences, Hopkinton, Mass.). The separation of product and substrate was optimized by choosing voltages and pressure using Caliper's Optimizer software (Hopkinton, Mass.). The separation conditions used a downstream voltage of -500V, an upstream voltage of -2150V, and a screening pressure of -1.2 psi. The product and substrate fluorophore were excited at 488 nm and detected at 530 nm. Substrate conversion was calculated from the electropherogram using HTS Well Analyzer software (Caliper Life Sciences, Hopkinton, Mass.). Ki values for the test compound were calculated.
Measurement: Ki = 0.032 nM (pChEMBL 10.489999771118164)
Descriptors: MW 412.46, ALogP 1.03, HBA 9, HBD 2, PSA 104.88, QED 0.48
Compound (SMILES): COc1cc(C2=C(c3cn(CCCN(C)C)c4ccccc34)C(=O)NC2=O)cc(OC)c1OC
InChIKey: PAIIKMRDIHPUKN-UHFFFAOYSA-N
Target: Glycogen synthase kinase-3 beta (SINGLE PROTEIN), Homo sapiens
UniProt: P49841
Assay: Kinase Inhibition Assay: The effect of test compounds on the activity of the protein kinases GSK-3β, VEGFR-2 and FLT-3 was evaluated based on half maximal inhibitory concentration (IC50) values determined by Millipore UK Ltd; Gemini Crescent; Dundee Technology Park; Dundee DD2 1SW; UK (IC50Profiler). Detailed protocols can be found at: www.millipore.com/drugdiscovery/dd3/assayprotocols.
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 463.53, ALogP 3.19, HBA 7, HBD 1, PSA 82.03, QED 0.49