lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): O=C(NCc1cccs1)c1cnc2c(SSc3cccc4cc(C(=O)NCc5cccs5)cnc34)cccc2c1
InChIKey: AEWRJZQAZXRVBK-UHFFFAOYSA-N
Target: STAM-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: O95630
Assay: Inhibition of AMSH (unknown origin) using DiUbK63TAMRA as substrate by fluorescence assay
Measurement: IC50 = 1300 nM (pChEMBL 5.889999866485596)
Descriptors: MW 598.8, ALogP 7.57, HBA 8, HBD 2, PSA 83.98, QED 0.17
Compound (SMILES): Cc1ccc(C(=O)NCC2NCCc3ccccc32)cc1C
InChIKey: VWJLMWIMGWFRII-UHFFFAOYSA-N
Target: Kappa-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41145
Assay: Agonist activity at human KOR expressed in HEK293 cells assessed as inhibition of forskolin-induced adenylyl cyclase-mediated cAMP accumulation after 30 mins by HTRF assay
Measurement: IC50 = 8930 nM (pChEMBL 5.050000190734863)
Descriptors: MW 294.4, ALogP 2.92, HBA 2, HBD 2, PSA 41.13, QED 0.91
Compound (SMILES): Cc1c(C#N)c(=O)oc2cc(OCc3ccc(Br)cc3)ccc12
InChIKey: BIGMVPGNLCXHRW-UHFFFAOYSA-N
Target: Cholinesterase (SINGLE PROTEIN), Equus caballus
UniProt: P81908
Assay: Inhibition of equine serum BuChE using S-butylthiocholine iodide as substrate preincubated for 10 mins followed by substrate addition measured after 10 mins by Ellman's method
Measurement: IC50 > 100000 nM (pChEMBL 4)
Descriptors: MW 370.2, ALogP 4.31, HBA 4, HBD 0, PSA 63.23, QED 0.64
Compound (SMILES): O=C(NO)c1cccc(-c2csc(CSc3ccccc3)n2)c1
InChIKey: KWDXMPXINALNFB-UHFFFAOYSA-N
Target: Histone deacetylase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P56524
Assay: Inhibition of HDAC4 (unknown origin) by fluorescence assay
Measurement: IC50 > 100000 nM (pChEMBL 4)
Descriptors: MW 342.45, ALogP 4.22, HBA 5, HBD 2, PSA 62.22, QED 0.41
Compound (SMILES): COc1cc(CNC(=O)C#Cc2cn(Cc3ccc(C(=O)NCCO)cc3)c(=O)[nH]c2=O)ccc1F
InChIKey: AZHGXAJGYOMYKJ-UHFFFAOYSA-N
Target: Collagenase 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P45452
Assay: Inhibition of C-terminal tagged human recombinant MMP13 catalytic domain expressed in Escherichia coli using (5-FAM/QX) FRET peptide as substrate by fluorescence assay
Measurement: IC50 = 72.44 nM (pChEMBL 7.139999866485596)
Descriptors: MW 494.48, ALogP 0.12, HBA 7, HBD 4, PSA 142.52, QED 0.33
Compound (SMILES): C[C@@]1(O)CC[C@H]2[C@H](CC[C@@H]3[C@@H]2CC[C@]2(C)[C@@H](C(=O)Cn4cc(C#N)cn4)CC[C@@H]32)C1
InChIKey: HARRKNSQXBRBGZ-GVKWWOCJSA-N
Target: Cytochrome P450 2C19 (SINGLE PROTEIN), Homo sapiens
UniProt: P33261
Assay: Inhibition of CYP2C19 (unknown origin)
Measurement: IC50 > 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 409.57, ALogP 4.34, HBA 5, HBD 1, PSA 78.91, QED 0.81
Compound (SMILES): Cc1cc(COc2ccc(S(=O)(=O)N[C@@H]3COC[C@@H]3C(=O)NO)cc2)c2ccccc2n1
InChIKey: STQWVQNLHJHEFT-PZJWPPBQSA-N
Target: Disintegrin and metalloproteinase domain-containing protein 17 (SINGLE PROTEIN), Mus musculus
UniProt: Q9Z0F8
Assay: Inhibition of TACE in mouse RAW264.7 cells assessed as inhibition of LPS-stimulated TNFalpha production pretreated for 1 hr followed by LPS stimulation for 4 hrs by ELISA
Measurement: IC50 = 12.59 nM (pChEMBL 7.900000095367432)
Descriptors: MW 457.51, ALogP 1.92, HBA 7, HBD 3, PSA 126.85, QED 0.37
Compound (SMILES): CCOC(=O)[C@@H]1C[C@@H]1Cc1c(Br)c(Br)c(OC)c(OC)c1Br
InChIKey: HBSUSNOAQHGZPQ-HTQZYQBOSA-N
Target: Carbonic anhydrase 12 (SINGLE PROTEIN), Homo sapiens
UniProt: O43570
Assay: Inhibition of human CA12 by stopped-flow CO2 hydration method
Measurement: Ki = 6600 nM (pChEMBL 5.179999828338623)
Descriptors: MW 501.01, ALogP 4.73, HBA 4, HBD 0, PSA 44.76, QED 0.42
Compound (SMILES): Cc1nncn1C1CCN(Cc2cc3nc(-c4cccc5[nH]ccc45)nc(N4CCOCC4)c3s2)CC1
InChIKey: DALQHZZOEYFELJ-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: Inhibition of PI3Kdelta (unknown origin) using PIP2 as substrate preincubated for 15 mins followed by substrate addition measured after 60 mins in presence of ATP by HTRF method
Measurement: IC50 = 3.981 nM (pChEMBL 8.399999618530273)
Descriptors: MW 514.66, ALogP 4.41, HBA 9, HBD 1, PSA 87.99, QED 0.37
Compound (SMILES): CCCCCCCCCCCC(=O)OCC(COP(=O)(O)OCCNC(=O)c1ccc(Cn2c(=O)[nH]c3c(N)nc(OCCOC)nc32)cc1)OC(=O)CCCCCCCCCCC
InChIKey: DFFGHPOFYMBXJP-UHFFFAOYSA-N
Target: Cytochrome P450 2D6 (SINGLE PROTEIN), Homo sapiens
UniProt: P10635
Assay: Fluorescent High Throughput P450 Assays: The interaction of SC12 with cytochrome P450 enzymes was tested using Fluorescent High Throughput P450 assays (Gentest); The IC50s of the compounds was calculated on isoenzymes (CYP1A2, CYP2C9, CYP2C19, CYP2C8, CYP2B6, CYP2D6, CYP2E1, CYP3A4 and CYP3A5).Materials and Methods: Inhibition of the P450 isoforms was measured in specific assays using specific substrates that become fluorescent upon CYP metabolism. Compounds, dissolved in ACN (acetonitrile) (CYP2E1, CYP2C8, CYP2B6, CYP3A5) or DMSO (all remaining isoforms), were tested in duplicate (n=2) in concentration-response curves (eight concentrations) in a 96-well plate containing incubation/NADPH regenerating buffer. Specific isoenzymes and substrates were added and incubated at 37° C. Reactions were terminated at different times, depending on the assays, and plates read on a Fluoroskan Ascent at the appropriate emission/excitation wavelengths.
Measurement: IC50 = 67000 nM (pChEMBL 4.170000076293945)
Descriptors: MW 921.08, ALogP 7.94, HBA 15, HBD 4, PSA 245.51, QED 0.02
Compound (SMILES): O=C1NOC[C@H]1NC[C@H]1CC[C@H](c2nnn3cnc4[nH]ccc4c23)CC1
InChIKey: FJUXWOXQAQGZDM-NQBHXWOUSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
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 = 370 nM (pChEMBL 6.429999828338623)
Descriptors: MW 355.4, ALogP 0.9, HBA 7, HBD 3, PSA 109.23, QED 0.64
Compound (SMILES): COC(=O)c1ccc2c(c1)-c1cccc(c1)[C@@H](N1CCC(c3c(F)ccc(Cl)c3F)=CC1=O)CCC[C@@H](C)C(=O)N2
InChIKey: PQMIPRJGMKRLRT-CLYVBNDRSA-N
Target: Plasma kallikrein (SINGLE PROTEIN), Homo sapiens
UniProt: P03952
Assay: In Vitro Assay: Plasma kallikrein determinations were made in 0.1 M sodium phosphate buffer at a pH of 7.5 containing 0.1-0.2 M sodium chloride and 0.5% PEG 8000. Determinations were made using purified human plasma kallikrein (Enzyme Research Laboratories) at a final assay concentration of 200 pM and the synthetic substrate S-2302 (H-(D)-Pro-Phe-Arg-pNA; CHROMOGENIX®) at a concentration of 0.00008-0.0004 M.
Measurement: Ki = 1398 nM (pChEMBL 5.849999904632568)
Descriptors: MW 579.04, ALogP 7.19, HBA 4, HBD 1, PSA 75.71, QED 0.26
Compound (SMILES): COc1ccc2c3c1O[C@H]1[C@@]4(OC)CC[C@@]5(C[C@@H]4COCc4ccccc4)[C@@H](C2)N(CC(N)=O)CC[C@]315
InChIKey: IFEHHNGEBPOACA-LDGIGSNBSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: Functional Assay: [35S]GTPgammaS functional assays were conducted using freshly thawed u-receptor membranes (Perkin Elmer, Shelton, Conn.). Assay reactions were prepared by sequentially adding the following reagents to binding buffer (100 mM NaCl, 10 mM MgCl2, 20 mM HEPES, pH 7.4) on ice (final concentrations indicated): membrane protein (0.026 mg/mL), saponin (10 mg/mL), GDP (3 mM) and [35S]GTPgammaS (0.20 nM; Perkin Elmer, Shelton, Conn.). The prepared membrane solution (190 ul/well) was transferred to 96-shallow well polypropylene plates containing 10 ul of 20x concentrated stock solutions of the agonist [D-Ala2, N-methyl-Phe4 Gly-ol5]-enkephalin (DAMGO) prepared in dimethyl sulfoxide (DMSO). Plates were incubated for 30 min at about 25° C. with shaking. Reactions were terminated by rapid filtration onto 96-well Unifilter GF/B filter plates (Perkin Elmer, Shelton, Conn.) using a 96-well tissue harvester (Perkin Elmer, Shelton, Conn.) followed by three filtration washes with 2001 of ice-cold binding buffer.
Measurement: EC50 = 874.31 nM (pChEMBL 6.059999942779541)
Descriptors: MW 504.63, ALogP 3.21, HBA 6, HBD 1, PSA 83.25, QED 0.59
Compound (SMILES): COC(=O)Nc1ccc2c(c1)NC(=O)[C@H](C)CCC[C@H](N1CCC(c3cccc(OC)c3)=CC1=O)c1cc-2ccn1
InChIKey: BURCJSBKPJCHNP-OLILMLBXSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: Inhibition Assay: Factor XIa determinations were made in 50 mM HEPES buffer at pH 7.4 containing 145 mM NaCl, 5 mM KCl, and 0.1% PEG 8000 (polyethylene glycol; JT Baker or Fisher Scientific). Determinations were made using purified human Factor XIa at a final concentration of 25-200 pM (Haematologic Technologies) and the synthetic substrate S-2366 (pyroGlu-Pro-Arg-pNA; CHROMOGENIX® or AnaSpec) at a concentration of 0.0002-0.001 M.
Measurement: Ki > 424 nM (pChEMBL 6.373000144958496)
Descriptors: MW 554.65, ALogP 6.05, HBA 6, HBD 2, PSA 109.86, QED 0.41
Compound (SMILES): COCCOCCOCCOCCO[C@H]1CC[C@@]2(O)C3Cc4ccc(O)c5c4[C@@]2(CCN3C)[C@H]1O5
InChIKey: PNZNYWUSPRKEKE-TZNCDMKYSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: HiRange Homogenous Time-Resolved Fluorescence (HTRF) Assay: Briefly, suspensions of cells expressing either the mu, kappa or delta opioid receptors were prepared in buffer containing 0.5 mM isobutyl-methyl xanthine (IBMX). Cells were incubated with varying concentrations of PEG-opioid conjugates and 3 uM forskolin for 30 minutes at room temperature. cAMP was detected following a two-step assay protocol per the manufacturer's instructions and time resolved fluorescence was measured with the following settings: 330 nm excitation; 620 nm and 665 nm emission; 380 nm dichroic mirror. The 665 nm/620 nm ratio is expressed as Delta F % and test compound-related data is expressed as a percentage of average maximum response in wells without forskolin. EC50 values were calculated for each compound from a sigmoidal dose-response plot of concentrations versus maximum response.
Measurement: EC50 = 128 nM (pChEMBL 6.889999866485596)
Descriptors: MW 493.6, ALogP 1.26, HBA 9, HBD 2, PSA 99.08, QED 0.39
Compound (SMILES): COc1ccc2c3c1O[C@H]1[C@@]4(OC)CC[C@@]5(C[C@@H]4CSc4nc6ccccc6s4)[C@@H](C2)N(CC2CC2)CC[C@]315
InChIKey: FSKVJNNDFYMKCF-QWEWFXQASA-N
Target: Kappa-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41145
Assay: Functional Assay: Functional [35S]GTPgammaS binding assays were conducted as follows. kippa opioid receptor membrane solution was prepared by sequentially adding final concentrations of 0.026 ug/ul kippa membrane protein (in-house), 10 ug/mL saponin, 3 uM GDP and 0.20 nM [35S]GTPgammaS to binding buffer (100 mM NaCl, 10 mM MgCl2, 20 mM HEPES, pH 7.4) on ice. The prepared membrane solution (190 ul/well) was transferred to 96-shallow well polypropylene plates containing 10 ul of 20x concentrated stock solutions of agonist prepared in DMSO. Plates were incubated for 30 min at a temperature of about 25° C. with shaking. Reactions were terminated by rapid filtration onto 96-well Unifilter GF/B filter plates (Perkin Elmer, Shelton, Conn.) using a 96-well tissue harvester (Packard) and followed by three filtration washes with 2001 ice-cold binding buffer (10 mM NaH2PO4, 10 mM Na2HPO4, pH 7.4). Filter plates were subsequently dried at 50° C. for 2-3 hours.
Measurement: EC50 = 63.79 nM (pChEMBL 7.199999809265137)
Descriptors: MW 560.79, ALogP 6.32, HBA 7, HBD 0, PSA 43.82, QED 0.32
Compound (SMILES): CN(C)[C@@H]1CCN(C[C@H]2CC[C@H](c3nnn4cnc5[nH]ccc5c34)CC2)C1
InChIKey: UHIYATACPWKMNN-BZUAXINKSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
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 367.5, ALogP 2.52, HBA 6, HBD 1, PSA 65.35, QED 0.77
Compound (SMILES): CCOC(=O)OC(C(C)C)n1nnc(-c2c(-c3ccc(C(=O)N(c4ncccc4C)[C@@H]4CCCNC4)cc3)cnn2C)n1
InChIKey: XMCIKMQAUXKSRZ-WIZGVZBGSA-N
Target: Proprotein convertase subtilisin/kexin type 9 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8NBP7
Assay: Cell Free Assay: In order to eliminate the permeability barrier inherent to the WT7 and SCHH cell-based assays a cell-free system was also established to access compound activity. A sequence containing the full length human PCSK9 (NCBI reference identifier, NM-174936.3, where coding sequence start annotated at position 363) along with 84 additional 3â¿² nucleotides, comprising a V5 tag and polylinkinker followed by an in frame modified firefly luciferase reporter (corresponding to nucleotide positions 283-1929 of pGL3, Gen Bank reference identifier JN542721.1) was cloned into the pT7CFE1 expression vector (ThermoScientific). The construct was then in-vitro transcribed using the MEGAscript T7 Kit (Life Technologies) and RNA subsequently purified incorporating the MEGAclear Kit (Life Technologies) according to manufacturer's protocols. HeLa cell lysates were prepared following the protocols described by Mikami (reference is Cell-Free Protein Synthesis Systems with Extracts from Cultured Human Cells, S. Mikami, T. Kobayashi and H. Imataka; from Methods in Molecular Biology, vol. 607, pages 43-52, Y. Endo et al. (eds.), Humana Press, 2010) with the following modifications.
Measurement: IC50 = 6200 nM (pChEMBL 5.210000038146973)
Descriptors: MW 587.69, ALogP 4.17, HBA 12, HBD 1, PSA 142.18, QED 0.28
Compound (SMILES): C[C@]1(c2nc(NC(=O)c3ncccc3Cl)ccc2F)N=C(N)OCC1(F)F
InChIKey: MQFTZDVMSMFRAU-OAHLLOKOSA-N
Target: Amyloid-beta precursor protein (SINGLE PROTEIN), Homo sapiens
UniProt: P05067
Assay: Inhibition Assay: Chinese hamster ovary cells are transfected with the human gene for amyloid precursor protein. The cells are plated at a density of 8000 cells/well into 96-well microtiter plates and cultivated for 24 hours in DMEM cell culture medium containing 10% FCS. The test compound is added to the cells at various concentrations, and the cells are cultivated for 24 hours in the presence of the test compound. The supernatants are collected, and the concentration of amyloid peptide 1-40 is determined using state of the art immunoassay techniques, for example sandwich ELISA, homogenous time-resolved fluorescence (HTRF) immunoassay, or electro-chemiluminescence immunoassay. The potency of the compound is calculated from the percentage of inhibition of amyloid peptide release as a function of the test compound concentration.
Measurement: IC50 = 88 nM (pChEMBL 7.059999942779541)
Descriptors: MW 399.76, ALogP 2.72, HBA 6, HBD 2, PSA 102.49, QED 0.83
Compound (SMILES): COc1ccc2c(c1)[C@]13CCN(CC4CC4)[C@H](C2)[C@]1(O)C[C@H](CCOCc1ccccc1)C(=O)C3
InChIKey: KYHDFAGHHSCKRP-PBNZBDRISA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: [35S]GTPγS Functional Assay: [35S]GTPγS functional assays were conducted using freshly thawed μ-receptor membranes prepared from a cell line expressing recombinant μ opioid receptor in a HEK-293, CHO or U-2 OS cell background or purchased from a commercial source (Perkin Elmer, Shelton, Conn.; or DiscovRx, Fremont, Calif.). Assay reactions were prepared by sequentially adding the following reagents to binding buffer (100 mM NaCl, 10 mM MgCl2, 20 mM HEPES, pH 7.4) on ice (final concentrations indicated): membrane protein (0.026 mg/mL), saponin (10 mg/mL), GDP (3 mM) and [35S]GTPγS (0.20 nM; Perkin Elmer, Shelton, Conn.). The prepared membrane solution (190 μl/well) was transferred to 96-shallow well polypropylene plates containing 10 μl of 20× concentrated stock solutions of the agonist [D-Ala2, N-methyl-Phe4 Gly-ol5]-enkephalin (DAMGO) prepared in dimethyl sulfoxide (DMSO). Plates were incubated for 30 min at about 25° C. with shaking. Reactions were terminated by rapid filtration onto 96-well Unifilter GF/B filter plates (Perkin Elmer, Shelton, Conn.) using a 96-well tissue harvester (Perkin Elmer, Shelton, Conn.) followed by three filtration washes with 200 μl of ice-cold wash buffer (10 mM NaH2PO4, 10 mM Na2HPO4, pH 7.4). Filter plates were subsequently dried at 50° C. for 2-3 hours. BetaScint scintillation cocktail (Perkin Elmer, Shelton, Conn.) was added (50 μl/well) and plates were counted using a Packard Top-Count for 1 min/well.
Measurement: EC50 = 38.6 nM (pChEMBL 7.409999847412109)
Descriptors: MW 475.63, ALogP 4.29, HBA 5, HBD 1, PSA 59, QED 0.58
Compound (SMILES): CC(Nc1ncc(F)c(N2C(=O)OC[C@@H]2[C@@H](C)O)n1)c1nc(-c2ccc(Cl)c(OC(F)(F)F)c2)no1
InChIKey: WXPAQCJZFINGNQ-VPQWPZGESA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Mutant IDH1 Biochemical Assay: LC-MS Detection of 2-HG: Mutant IDH1 R132H catalytic activity was monitored using the quantitative liquid chromatography/mass spectrometry (LC-MS) detection of 2-HG, a product of the NADPH-dependent alpha-KG reduction reaction.More specifically, the biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Costar, Cat. No. 781201) using a final reaction volume of 30 μL and the following assay buffer conditions: 50 mM HEPES pH 7.4, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.02% BSA, 5 uM NADPH and 100 uM alpha-KG.The final reaction mixture contained 3.3% DMSO and inhibitors with concentrations ranging 0.02-50 μM. The IDH1 enzyme was used at a final concentration of 0.25 nM. Following 45 minutes incubation, the reaction mixtures were quenched by the addition of 10 μL of 16% formic acid containing 800 nM of 5-carbon labeled 13C-2-HG). The protein was then precipitated by the addition of 2.5 volumes of acetonitrile followed by centrifugation (3000×g, 20 minutes). The concentration of 2-HG in the resulting supernatants was measured by LC-MS (see below).LC-MS method. Reaction mixture supernatants were submitted to chromatographic separation on a BiobasicAX column (2.1 mm×20 mm, 5 μm particle, Thermo Scientific Inc.). The chromatographic mobile phases were A) 25 mM ammonium biocarbonate and B) acetonitrile (0.1% ammonium hydroxide). Nicotinamide was eluted at 1 ml/min using a 85-5% B gradient over 0.9 minutes (Agilent 1200SL LC system, Thermofisher LX-4 autosampler) and analyzed by multiple reaction monitoring (MRM) on a API4000 QTrap mass spectrometer (ABSciex, Framingham, Mass.) in the positive electrospray ionization (ESI+) mode. The mass transition for 2-HG and 13C-2-HG were 147→129 and 152→134, respectively. The relative responses (2-HG/13C-2-HG) were measured at varied inhibitor concentrations and used to calculate inhibitory IC50 values (normalized IC50 regression curves).
Measurement: IC50 = 77 nM (pChEMBL 7.110000133514404)
Descriptors: MW 532.84, ALogP 4.1, HBA 10, HBD 2, PSA 135.73, QED 0.43
Compound (SMILES): N[C@@H]1CCCN(c2cccc(-n3ncc4cnc(-c5cncs5)cc43)n2)C1
InChIKey: BQVZFNYKORVNKX-CQSZACIVSA-N
Target: Serine/threonine-protein kinase pim-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P11309
Assay: In Vitro Cell Proliferation Potency Assay: BaF3, a murine interleukin-3 dependent pro-B cell line, parental cells, BaF3 PIM1 cells, BaF3 PIM2 cells, and MM1.S (multiple myeloma) cells were seeded at 2 k/well, 5 k/well, 5 k/well, and 10 k/well respectively, in a 384-well plate, at 45 μL/well. Test compound was added at 5 μL/well. BaF3 cells (parental and transfected) were incubated overnight, while MM1.S cells were incubated for 72 hours at 37° C., 5% CO2. CELL TITER GLO® Reagent (Promega) was added at 50 μL/well, the plates were incubated for 30 minutes, and their luminescence read on an HT Analyst.
Measurement: IC50 = 1700 nM (pChEMBL 5.769999980926514)
Descriptors: MW 377.48, ALogP 2.87, HBA 8, HBD 1, PSA 85.75, QED 0.59
Compound (SMILES): O=C(Nc1ccc(SC(F)(F)Cl)cc1)c1cnc(N2CC[C@@H](O)C2)c(-c2cncc(F)c2)c1
InChIKey: XDWCEJMZRFFPOS-QGZVFWFLSA-N
Target: Tyrosine-protein kinase ABL1 (SINGLE PROTEIN), Homo sapiens
UniProt: P00519
Assay: Cellular Proliferation Assay: Test compounds were resuspended in DMSO at a concentration of 10 mM. A serial three-fold dilution of each compound with DMSO was performed in 384-well plates using the Janus Liquid Dispenser (PerkinElmer). Compound was delivered to the assay plates containing 2500 cells in a 50 μL volume via Acoustic delivery from an ATS-100 (EDC). For Ba/F3-BCR-ABL1-WT cell assays, 2 nL of each compound dilution was transferred to the assay plate for final assay concentrations of 0.4 μM, 0.13 μM, 0.044 μM, 0.015 μM, 0.005 μM, 0.001 μM, 0.00033 μM, 0.00011 μM, 0.000037 μM, 0.000012 μM. For Ba/F3-WT and Ba/F3-BCR-ABL1-T315I cell assays, 50 nL of each compound dilution was transferred to the assay plate for final assay concentrations of 10 μM, 3.33 μM, 1.11 μM, 0.37 μM, 0.12 μM, 0.041 μM, 0.014 μM, 0.0046 μM, 0.0015 μM, 0.00051 μM.Cells were incubated at 37° C. in a humidified environment with 5% car
Measurement: IC50 = 10 nM (pChEMBL 8)
Descriptors: MW 494.93, ALogP 4.99, HBA 6, HBD 2, PSA 78.35, QED 0.37
Compound (SMILES): CC#CC(=O)N1CCC[C@@H](n2nc(-c3ccc(Oc4ccccc4)cc3)c3c(N)ncnc32)C1
InChIKey: KEOXYNOFRLIAAM-LJQANCHMSA-N
Target: Epidermal growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P00533
Assay: HotSpot kinase assay: IC50s were determined using the in vitro HotSpot kinase assay (purified enzymes, 33P-ATP, an appropriate substrate and 1 μM ATP.). For enzyme inhibition assays, compounds were tested in range of ten concentrations from 10 uM to 0.0005 uM using purified enzymes and the Hotspot kinase assay. Reaction conditions were 1 uM ATP, one hour incubation with inhibitor, and kinase activity detected using 33-ATP phosphorylation of an appropriately selected peptide substrate.
Measurement: IC50 = 435.3 nM (pChEMBL 6.360000133514404)
Descriptors: MW 452.52, ALogP 4.05, HBA 7, HBD 1, PSA 99.16, QED 0.47
Compound (SMILES): C[C@@H]1CCC[C@H](N2CCC(c3c(-c4cn[nH]c4)ccc(Cl)c3F)=CC2=O)c2cc(ccn2)-c2ccccc2NC1=O
InChIKey: GWFIQOMARMQBJV-GDJIYFAZSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: Inhibition Assay: Factor XIa determinations were made in 50 mM HEPES buffer at pH 7.4 containing 145 mM NaCl, 5 mM KCl, and 0.1% PEG 8000 (polyethylene glycol; JT Baker or Fisher Scientific). Determinations were made using purified human Factor XIa at a final concentration of 25-200 pM (Haematologic Technologies) and the synthetic substrate S-2366 (pyroGlu-Pro-Arg-pNA; CHROMOGENIX® or AnaSpec) at a concentration of 0.0002-0.001 M.
Measurement: Ki = 2.73 nM (pChEMBL 8.5600004196167)
Descriptors: MW 570.07, ALogP 7.05, HBA 4, HBD 2, PSA 90.98, QED 0.28
Compound (SMILES): CC(C)[C@@H](C(=O)N1CCC(Nc2ncc(C(=O)NO)cn2)(c2ccccc2)CC1)c1ccccc1
InChIKey: DOSFADKGXFNINV-HSZRJFAPSA-N
Target: Protein deacetylase HDAC6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UBN7
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 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 473.58, ALogP 3.97, HBA 6, HBD 3, PSA 107.45, QED 0.35
Compound (SMILES): COc1cc(C(O)P(=O)(O)CC[C@@H](N)C(=O)O)ccc1OCC(=O)O
InChIKey: LNLCIHWIBXKXQU-UCWRFOARSA-N
Target: Metabotropic glutamate receptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P35400
Assay: Pharmacological Assay: Metabotropic glutamate receptors were transiently transfected in HEK293 cells by electroporation as described elsewhere (Brabet I. et al., 1998) and plated in 96-well microplates. The high affinity glutamate transporter EAAC1 was co-transfected with the receptor in order to avoid any influence of glutamate released by the cells in the assay medium. In the experiments carried out by the inventors, Group-III mGluRs were co-transfected with a chimeric G-protein which couples the activation of the receptor to the phospholipase-C (PLC) pathway. Thus receptor activation induces production of inositol phosphate (IP) which in turn induces intracellular Ca2+ release. Receptor activity was then determined by measurement of the IP production or Ca release as already described (Goudet C. et al., PNAS 2004). For intracellular calcium measurements, cells expressing mGluRs were loaded with Ca2+-sensitive fluorescent dye Fluo-4 AM (Invitrogen, Cergy-Pontoise, France) dissolved in Hanks' balanced Salt Solution.
Measurement: EC50 = 11500 nM (pChEMBL 4.940000057220459)
Descriptors: MW 377.29, ALogP 0.22, HBA 7, HBD 5, PSA 176.61, QED 0.35
Compound (SMILES): CCOCc1c(-c2cc3cc(C)cc(OC)c3s2)c2c(N)ncnn2c1CN[C@@H]1CCNC1=O
InChIKey: BQGCDWWKWBZGAX-MRXNPFEDSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Kinase Assay: For the assay, 50 nl of each stock 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 the above FGFR-3 fusion protein in aqueous assay buffer [8 mM MOPS pH 7.0, 10 mM magnesium acetate, 1.0 mM dithiothreitol, 0.05% (w/v) bovine serum albumin (BSA), 0.07% (v/v) Tween-20, 0.2 mM EDTA] was added, and the mixture was incubated for 15 min at 22° C. to allow pre-binding of the test compound to the enzyme. Then, the kinase reaction was started by the addition of 3 ul of a solution of adenosine triphosphate (ATP, 16.7 uM; final concentration in the 5 ul assay volume=10 uM) and substrate (0.8 uM; final concentration in the 5 ul assay volume=0.5 uM) in assay buffer, and the resulting mixture was incubated for a reaction time of 60 min at 22° C. The concentration of FGFR-3 fusion protein was adjusted depending on the activity of the enzyme lot.
Measurement: IC50 = 17.7 nM (pChEMBL 7.75)
Descriptors: MW 480.59, ALogP 3.02, HBA 9, HBD 3, PSA 115.8, QED 0.35
Compound (SMILES): CC(Nc1nccc(N2C(=O)OC[C@@H]2[C@@H](C)O)n1)c1cnc(-c2ccccc2)s1
InChIKey: JBIBAWVBYNZUKK-JYRZLJSNSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Mutant IDH1 Biochemical Assay: LC-MS Detection of 2-HG: Mutant IDH1 R132H catalytic activity was monitored using the quantitative liquid chromatography/mass spectrometry (LC-MS) detection of 2-HG, a product of the NADPH-dependent alpha-KG reduction reaction.More specifically, the biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Costar, Cat. No. 781201) using a final reaction volume of 30 μL and the following assay buffer conditions: 50 mM HEPES pH 7.4, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.02% BSA, 5 uM NADPH and 100 uM alpha-KG.The final reaction mixture contained 3.3% DMSO and inhibitors with concentrations ranging 0.02-50 μM. The IDH1 enzyme was used at a final concentration of 0.25 nM. Following 45 minutes incubation, the reaction mixtures were quenched by the addition of 10 μL of 16% formic acid containing 800 nM of 5-carbon labeled 13C-2-HG). The protein was then precipitated by the addition of 2.5 volumes of acetonitrile followed by centrifugation (3000×g, 20 minutes). The concentration of 2-HG in the resulting supernatants was measured by LC-MS (see below).LC-MS method. Reaction mixture supernatants were submitted to chromatographic separation on a BiobasicAX column (2.1 mm×20 mm, 5 μm particle, Thermo Scientific Inc.). The chromatographic mobile phases were A) 25 mM ammonium biocarbonate and B) acetonitrile (0.1% ammonium hydroxide). Nicotinamide was eluted at 1 ml/min using a 85-5% B gradient over 0.9 minutes (Agilent 1200SL LC system, Thermofisher LX-4 autosampler) and analyzed by multiple reaction monitoring (MRM) on a API4000 QTrap mass spectrometer (ABSciex, Framingham, Mass.) in the positive electrospray ionization (ESI+) mode. The mass transition for 2-HG and 13C-2-HG were 147→129 and 152→134, respectively. The relative responses (2-HG/13C-2-HG) were measured at varied inhibitor concentrations and used to calculate inhibitory IC50 values (normalized IC50 regression curves).
Measurement: IC50 = 4032 nM (pChEMBL 5.389999866485596)
Descriptors: MW 411.49, ALogP 3.48, HBA 8, HBD 2, PSA 100.47, QED 0.64
Compound (SMILES): CCCCCCCC(F)(F)CCCCCC/C=C/[C@H](C(O)N[C@@H](Cc1ccc(OCCCC)cc1)C(=O)O)[C@@](O)(CCOC)C(=O)O
InChIKey: OWMVKQWKXXHJGW-HNOZPZLJSA-N
Target: Cytochrome P450 2C9 (SINGLE PROTEIN), Homo sapiens
UniProt: P11712
Assay: CYP Inhibition Assay (With Preincubation): With Preincubation: To a 96 deep well plate was added 2.5 uL of the test substance solution (x200 concentration), and 442.5 uL of Incubation Mixture, and the solution was mixed and warmed to 37° C. for 5 minutes. To this incubation solution 50 uL of 10 mM NADPH was added and preincubated at 37° C. for 30 minutes. After the preincubation, 5 uL of the substrate solution was added and the solution was mixed and further incubated at 37° C. for 5 minutes for the reaction. After the reaction, 100 uL of the Incubation Mixture was removed, added to 150 uL of isopropanol containing IS and mixed. This mixture was transferred onto a multi-screen filter, centrifuged at 500xg at 4° C. for 10 minutes. The filtrate was used for LC-MS/MS analysis.
Measurement: IC50 = 24400 nM (pChEMBL 4.610000133514404)
Descriptors: MW 671.86, ALogP 7.13, HBA 7, HBD 5, PSA 145.55, QED 0.03
Compound (SMILES): CC(O)C#Cc1c(N)ncnc1N[C@H](C)c1nc2cccc(Cl)c2c(=O)n1-c1ccccc1
InChIKey: GLZTYCNWUIEVIB-YSSOQSIOSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P48736
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 > 1000 nM (pChEMBL 6)
Descriptors: MW 460.93, ALogP 3.32, HBA 8, HBD 3, PSA 118.95, QED 0.4
Compound (SMILES): COC(=O)c1cccc2c1-c1ccnc(c1)[C@@H](N1CCC(c3c(F)ccc(Cl)c3F)=CC1=O)CCC[C@@H](C)C(=O)N2
InChIKey: CXESAZISLSPVSR-NSYGIPOTSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: Inhibition Assay: Factor XIa determinations were made in 50 mM HEPES buffer at pH 7.4 containing 145 mM NaCl, 5 mM KCl, and 0.1% PEG 8000 (polyethylene glycol; JT Baker or Fisher Scientific). Determinations were made using purified human Factor XIa at a final concentration of 25-200 pM (Haematologic Technologies) and the synthetic substrate S-2366 (pyroGlu-Pro-Arg-pNA; CHROMOGENIX® or AnaSpec) at a concentration of 0.0002-0.001 M.
Measurement: Ki > 413 nM (pChEMBL 6.383999824523926)
Descriptors: MW 580.03, ALogP 6.58, HBA 5, HBD 1, PSA 88.6, QED 0.28
Compound (SMILES): N#Cc1cnc(NC(=O)N2CCCc3cc(CO)c(C=O)nc32)cc1O[C@@H]1CCOC1
InChIKey: JDSIDNMVOWOKBD-MRXNPFEDSA-N
Target: Fibroblast growth factor receptor 4 (SINGLE PROTEIN), Mus musculus
UniProt: Q03142
Assay: Inhibition of FGFR4 in mouse BAF3 cells assessed as decrease in FGFR4 phosphorylation incubated for 40 mins
Measurement: IC50 = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 423.43, ALogP 1.81, HBA 8, HBD 2, PSA 137.67, QED 0.69
Compound (SMILES): CC(Nc1nccc(N2C(=O)OC[C@@H]2[C@@H](C)O)n1)c1nc(-c2cccc(Cl)c2)no1
InChIKey: VDEXLVZLFHDDEG-MDLDFJCZSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Mutant IDH1 Biochemical Assay: LC-MS Detection of 2-HG: Mutant IDH1 R132H catalytic activity was monitored using the quantitative liquid chromatography/mass spectrometry (LC-MS) detection of 2-HG, a product of the NADPH-dependent alpha-KG reduction reaction.More specifically, the biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Costar, Cat. No. 781201) using a final reaction volume of 30 μL and the following assay buffer conditions: 50 mM HEPES pH 7.4, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.02% BSA, 5 uM NADPH and 100 uM alpha-KG.The final reaction mixture contained 3.3% DMSO and inhibitors with concentrations ranging 0.02-50 μM. The IDH1 enzyme was used at a final concentration of 0.25 nM. Following 45 minutes incubation, the reaction mixtures were quenched by the addition of 10 μL of 16% formic acid containing 800 nM of 5-carbon labeled 13C-2-HG). The protein was then precipitated by the addition of 2.5 volumes of acetonitrile followed by centrifugation (3000×g, 20 minutes). The concentration of 2-HG in the resulting supernatants was measured by LC-MS (see below).LC-MS method. Reaction mixture supernatants were submitted to chromatographic separation on a BiobasicAX column (2.1 mm×20 mm, 5 μm particle, Thermo Scientific Inc.). The chromatographic mobile phases were A) 25 mM ammonium biocarbonate and B) acetonitrile (0.1% ammonium hydroxide). Nicotinamide was eluted at 1 ml/min using a 85-5% B gradient over 0.9 minutes (Agilent 1200SL LC system, Thermofisher LX-4 autosampler) and analyzed by multiple reaction monitoring (MRM) on a API4000 QTrap mass spectrometer (ABSciex, Framingham, Mass.) in the positive electrospray ionization (ESI+) mode. The mass transition for 2-HG and 13C-2-HG were 147→129 and 152→134, respectively. The relative responses (2-HG/13C-2-HG) were measured at varied inhibitor concentrations and used to calculate inhibitory IC50 values (normalized IC50 regression curves).
Measurement: IC50 = 3977 nM (pChEMBL 5.400000095367432)
Descriptors: MW 430.85, ALogP 3.06, HBA 9, HBD 2, PSA 126.5, QED 0.61
Compound (SMILES): COCCOCCOCCOCCOCCOC[C@H]1C[C@@]23CC[C@]1(OC)[C@@H]1Oc4c(OC)ccc5c4[C@@]12CCN(CC1CC1)[C@@H]3C5
InChIKey: AOUNVUHADLIPSM-QTDSEMIKSA-N
Target: Kappa-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41145
Assay: Functional Assay: Functional [35S]GTPgammaS binding assays were conducted as follows. kippa opioid receptor membrane solution was prepared by sequentially adding final concentrations of 0.026 ug/ul kippa membrane protein (in-house), 10 ug/mL saponin, 3 uM GDP and 0.20 nM [35S]GTPgammaS to binding buffer (100 mM NaCl, 10 mM MgCl2, 20 mM HEPES, pH 7.4) on ice. The prepared membrane solution (190 ul/well) was transferred to 96-shallow well polypropylene plates containing 10 ul of 20x concentrated stock solutions of agonist prepared in DMSO. Plates were incubated for 30 min at a temperature of about 25° C. with shaking. Reactions were terminated by rapid filtration onto 96-well Unifilter GF/B filter plates (Perkin Elmer, Shelton, Conn.) using a 96-well tissue harvester (Packard) and followed by three filtration washes with 2001 ice-cold binding buffer (10 mM NaH2PO4, 10 mM Na2HPO4, pH 7.4). Filter plates were subsequently dried at 50° C. for 2-3 hours.
Measurement: EC50 = 816.86 nM (pChEMBL 6.090000152587891)
Descriptors: MW 645.83, ALogP 3.65, HBA 10, HBD 0, PSA 86.31, QED 0.18
Compound (SMILES): COC(=O)Nc1ccc2c(c1)NC(=O)[C@H](C)CCC[C@H](N1CCC(c3cc(Cl)ccc3C(F)(F)F)=CC1=O)c1cc-2ccn1
InChIKey: YGFGZLKRXRBORE-WRHNGFHOSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: Inhibition Assay: Factor XIa determinations were made in 50 mM HEPES buffer at pH 7.4 containing 145 mM NaCl, 5 mM KCl, and 0.1% PEG 8000 (polyethylene glycol; JT Baker or Fisher Scientific). Determinations were made using purified human Factor XIa at a final concentration of 25-200 pM (Haematologic Technologies) and the synthetic substrate S-2366 (pyroGlu-Pro-Arg-pNA; CHROMOGENIX® or AnaSpec) at a concentration of 0.0002-0.001 M.
Measurement: Ki = 0.09 nM (pChEMBL 10.050000190734863)
Descriptors: MW 627.06, ALogP 7.71, HBA 5, HBD 2, PSA 100.63, QED 0.31
Compound (SMILES): O=C(Nc1ccc(SC(F)(F)F)cc1)c1cnc(N2CC[C@@H](O)C2)c(-c2cncnc2)c1
InChIKey: UHBHXEYMLMAMQC-MRXNPFEDSA-N
Target: Tyrosine-protein kinase ABL1 (SINGLE PROTEIN), Homo sapiens
UniProt: P00519
Assay: Cellular Proliferation Assay: Test compounds were resuspended in DMSO at a concentration of 10 mM. A serial three-fold dilution of each compound with DMSO was performed in 384-well plates using the Janus Liquid Dispenser (PerkinElmer). Compound was delivered to the assay plates containing 2500 cells in a 50 μL volume via Acoustic delivery from an ATS-100 (EDC). For Ba/F3-BCR-ABL1-WT cell assays, 2 nL of each compound dilution was transferred to the assay plate for final assay concentrations of 0.4 μM, 0.13 μM, 0.044 μM, 0.015 μM, 0.005 μM, 0.001 μM, 0.00033 μM, 0.00011 μM, 0.000037 μM, 0.000012 μM. For Ba/F3-WT and Ba/F3-BCR-ABL1-T315I cell assays, 50 nL of each compound dilution was transferred to the assay plate for final assay concentrations of 10 μM, 3.33 μM, 1.11 μM, 0.37 μM, 0.12 μM, 0.041 μM, 0.014 μM, 0.0046 μM, 0.0015 μM, 0.00051 μM.Cells were incubated at 37° C. in a humidified environment with 5% car
Measurement: IC50 = 19 nM (pChEMBL 7.71999979019165)
Descriptors: MW 461.47, ALogP 3.97, HBA 7, HBD 2, PSA 91.24, QED 0.56
Compound (SMILES): COC(=O)c1cc2c(cc1N)NC(=O)[C@H](C)CCC[C@H](N1CCC(c3c(F)ccc(Cl)c3F)=CC1=O)c1cc-2ccn1
InChIKey: KZXNHUNTABHMLN-DXPJPUQTSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: Inhibition Assay: Factor XIa determinations were made in 50 mM HEPES buffer at pH 7.4 containing 145 mM NaCl, 5 mM KCl, and 0.1% PEG 8000 (polyethylene glycol; JT Baker or Fisher Scientific). Determinations were made using purified human Factor XIa at a final concentration of 25-200 pM (Haematologic Technologies) and the synthetic substrate S-2366 (pyroGlu-Pro-Arg-pNA; CHROMOGENIX® or AnaSpec) at a concentration of 0.0002-0.001 M.
Measurement: Ki = 4.54 nM (pChEMBL 8.34000015258789)
Descriptors: MW 595.05, ALogP 6.16, HBA 6, HBD 2, PSA 114.62, QED 0.21
Compound (SMILES): CO[C@]12CC[C@@]3(C[C@@H]1COCc1ccccc1)[C@H]1Cc4ccc(C(N)=O)c5c4[C@@]3(CCN1CC1CC1)[C@H]2O5
InChIKey: HQCCMURBRBVEDJ-ZZVPPEEFSA-N
Target: Kappa-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41145
Assay: Functional Assay: Functional [35S]GTPgammaS binding assays were conducted as follows. kippa opioid receptor membrane solution was prepared by sequentially adding final concentrations of 0.026 ug/ul kippa membrane protein (in-house), 10 ug/mL saponin, 3 uM GDP and 0.20 nM [35S]GTPgammaS to binding buffer (100 mM NaCl, 10 mM MgCl2, 20 mM HEPES, pH 7.4) on ice. The prepared membrane solution (190 ul/well) was transferred to 96-shallow well polypropylene plates containing 10 ul of 20x concentrated stock solutions of agonist prepared in DMSO. Plates were incubated for 30 min at a temperature of about 25° C. with shaking. Reactions were terminated by rapid filtration onto 96-well Unifilter GF/B filter plates (Perkin Elmer, Shelton, Conn.) using a 96-well tissue harvester (Packard) and followed by three filtration washes with 2001 ice-cold binding buffer (10 mM NaH2PO4, 10 mM Na2HPO4, pH 7.4). Filter plates were subsequently dried at 50° C. for 2-3 hours.
Measurement: EC50 = 0.7 nM (pChEMBL 9.149999618530273)
Descriptors: MW 514.67, ALogP 4.23, HBA 5, HBD 1, PSA 74.02, QED 0.57
Compound (SMILES): COC(=O)Nc1ccc2c(c1)NC(=O)[C@H](C)CC(=O)C[C@H](N1CCC(c3c(F)ccc(Cl)c3F)=CC1=O)c1cc-2ccn1
InChIKey: JMHUINMWXFHURP-DXPJPUQTSA-N
Target: Coagulation factor XI (SINGLE PROTEIN), Homo sapiens
UniProt: P03951
Assay: Inhibition Assay: Factor XIa determinations were made in 50 mM HEPES buffer at pH 7.4 containing 145 mM NaCl, 5 mM KCl, and 0.1% PEG 8000 (polyethylene glycol; JT Baker or Fisher Scientific). Determinations were made using purified human Factor XIa at a final concentration of 25-200 pM (Haematologic Technologies) and the synthetic substrate S-2366 (pyroGlu-Pro-Arg-pNA; CHROMOGENIX® or AnaSpec) at a concentration of 0.0002-0.001 M.
Measurement: Ki > 438 nM (pChEMBL 6.359000205993652)
Descriptors: MW 609.03, ALogP 6.15, HBA 6, HBD 2, PSA 117.7, QED 0.34
Compound (SMILES): CC(Nc1nccc(N2C(=O)OC[C@@H]2[C@@H](C)O)n1)c1nc(-c2ccc(Cl)c(C(F)(F)F)c2)no1
InChIKey: DNPZDRUUCRRSIU-WUMONGPASA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Mutant IDH1 Biochemical Assay: LC-MS Detection of 2-HG: Mutant IDH1 R132H catalytic activity was monitored using the quantitative liquid chromatography/mass spectrometry (LC-MS) detection of 2-HG, a product of the NADPH-dependent alpha-KG reduction reaction.More specifically, the biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Costar, Cat. No. 781201) using a final reaction volume of 30 μL and the following assay buffer conditions: 50 mM HEPES pH 7.4, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.02% BSA, 5 uM NADPH and 100 uM alpha-KG.The final reaction mixture contained 3.3% DMSO and inhibitors with concentrations ranging 0.02-50 μM. The IDH1 enzyme was used at a final concentration of 0.25 nM. Following 45 minutes incubation, the reaction mixtures were quenched by the addition of 10 μL of 16% formic acid containing 800 nM of 5-carbon labeled 13C-2-HG). The protein was then precipitated by the addition of 2.5 volumes of acetonitrile followed by centrifugation (3000×g, 20 minutes). The concentration of 2-HG in the resulting supernatants was measured by LC-MS (see below).LC-MS method. Reaction mixture supernatants were submitted to chromatographic separation on a BiobasicAX column (2.1 mm×20 mm, 5 μm particle, Thermo Scientific Inc.). The chromatographic mobile phases were A) 25 mM ammonium biocarbonate and B) acetonitrile (0.1% ammonium hydroxide). Nicotinamide was eluted at 1 ml/min using a 85-5% B gradient over 0.9 minutes (Agilent 1200SL LC system, Thermofisher LX-4 autosampler) and analyzed by multiple reaction monitoring (MRM) on a API4000 QTrap mass spectrometer (ABSciex, Framingham, Mass.) in the positive electrospray ionization (ESI+) mode. The mass transition for 2-HG and 13C-2-HG were 147→129 and 152→134, respectively. The relative responses (2-HG/13C-2-HG) were measured at varied inhibitor concentrations and used to calculate inhibitory IC50 values (normalized IC50 regression curves).
Measurement: IC50 = 1884 nM (pChEMBL 5.71999979019165)
Descriptors: MW 498.85, ALogP 4.08, HBA 9, HBD 2, PSA 126.5, QED 0.52
Compound (SMILES): Cc1nn(C)c(C)c1NS(=O)(=O)c1ccc(-c2cccc(-c3nccn3C)c2)cc1
InChIKey: PPRLWPOJFJTUNF-UHFFFAOYSA-N
Target: Glycylpeptide N-tetradecanoyltransferase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P30419
Assay: Enzyme Inhibition Assay: Measurements were performed using a modification of the scintillation proximity assay platform described previously by Georgopapadakou, N. H. et al. (22nd International Congress on Chemotherapy, 2001, Abstract P16.001). Compounds were solubilised in DMSO at a top concentration of 10 mM and serially diluted in half log steps to achieve a range of final assay concentrations of 100 uM to 1 nM. Compound at each concentration (100-fold final) was added to white 384 well plates in a volume of 0.5 ml. Human, A. fumigatus, T. brucei or L. major N-myristoyl transferase enzyme, dissolved to a working concentration of 10 nM in assay buffer (30 mM Tris/HCl pH 7.4, 0.5 mM EGTA, 0.5 mM EDTA, 1.25 mM DTT, 0.1% Triton X-100), was then added to columns 1 to 11 and 13 to 23 of the plates in a volume of 20 ml. To columns 12 and 24, 20 ml assay buffer was added to provide a no enzyme control. Following a 5 minute incubation at room temperature the substrates (GCGGSKVKPQPPQAK(Biotin)-Amide and myristoyl coenzyme A), dissolved in assay buffer, were added to all wells in a volume of 20 ml to start the reaction. The final concentrations of peptide and 3H-myristoyl coenzyme A were 0.5 mM and 125 nM respectively and the specific activity of the radiolabel was 8 Ci/mmol. Plates were then incubated at room temperature for up to 50 minutes (dependant upon the period of linearity for the different enzyme species) before SPA beads, suspended to 1 mg/ml in a stop solution (200 mM Phosphoric Acid/NaOH pH 4, 750 mM MgCl2), were added in a volume of 40 ml.
Measurement: IC50 = 2400 nM (pChEMBL 5.619999885559082)
Descriptors: MW 421.53, ALogP 3.91, HBA 6, HBD 1, PSA 81.81, QED 0.53
Compound (SMILES): CC(Nc1nccc(N2C(=O)OC[C@@H]2[C@@H](C)O)n1)c1nnc(-c2cccc(F)c2)o1
InChIKey: ILNTZEINCNMYCW-MDLDFJCZSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: LC-MS Detection of 2-HG Assay: The biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Costar, Cat. No. 781201) using a final reaction volume of 30 μL and the following assay buffer conditions: 50 mM HEPES pH 7.4, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.02% BSA, 5 uM NADPH and 100 uM alpha-KG. The final reaction mixture contained 3.3% DMSO and inhibitors with concentrations ranging 0.02-50 μM. The IDH1 enzyme was used at a final concentration of 0.25 nM. Following 45 minutes incubation, the reaction mixtures were quenched by the addition of 10 μL of 16% formic acid containing 800 nM of 5-carbon labeled 13C-2-HG). The protein was then precipitated by the addition of 2.5 volumes of acetonitrile followed by centrifugation (3000×g, minutes). The concentration of 2-HG in the resulting supernatants was measured by LC-MS (see below). LC-MS method. Reaction mixture supernatants were submitted to chromatographic separation on a BiobasicAX column (2.1 mm×20 mm, 5 μm particle, Thermo Scientific Inc.). The chromatographic mobile phases were A) 25 mM ammonium biocarbonate and B) acetonitrile (0.1% ammonium hydroxide). Nicotinamide was eluted at 1 ml/min using a 85-5% B gradient over 0.9 minutes (Agilent 1200SL LC system, Thermofisher LX-4 autosampler) and analyzed by multiple reaction monitoring (MRM) on a API4000 QTrap mass spectrometer (ABSciex, Framingham, Mass.) in the positive electrospray ionization (ESI+) mode. The mass transition for 2-HG and 13C-2-HG were 147→129 and 152→134, respectively.
Measurement: IC50 = 3511 nM (pChEMBL 5.460000038146973)
Descriptors: MW 414.4, ALogP 2.54, HBA 9, HBD 2, PSA 126.5, QED 0.63