lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Cc1nc(Nc2ncc(C(=O)Nc3c(C)cccc3Cl)s2)cc(N2CCN(CC(=O)NO)CC2)n1
InChIKey: XBSRSOMLIPRLGT-UHFFFAOYSA-N
Target: Proto-oncogene tyrosine-protein kinase Src (SINGLE PROTEIN), Homo sapiens
UniProt: P12931
Assay: Inhibition of human N-terminal GST-tagged full length human Src (Met1 to Leu536 residues) expressed in baculovirus expression system assessed as decrease in biotin signal transduction protein phosphorylation at Tyr160 preincubated for 5 mins followed by substrate addition measured after 30 mins by DELFIA method
Measurement: IC50 <= 100 nM (pChEMBL 7)
Descriptors: MW 517.02, ALogP 2.83, HBA 10, HBD 4, PSA 135.61, QED 0.28
Compound (SMILES): Cc1ccc(C2=C(C(=O)Nc3ccc(C)c(F)c3)C(=O)NC(c3ccc(OCCCC(F)(F)F)cc3)(C(F)(F)F)C2)cc1
InChIKey: LZPIKVIZUSUYBL-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 = 1465 nM (pChEMBL 5.829999923706055)
Descriptors: MW 608.55, ALogP 7.53, HBA 3, HBD 2, PSA 67.43, QED 0.16
Compound (SMILES): O=S1(=O)c2ccccc2CN1c1cc(F)cc(Oc2ccccc2)c1
InChIKey: PCSGJZXEGRHDSH-UHFFFAOYSA-N
Target: Free fatty acid receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5NUL3
Assay: Agonist activity at human FFA4 expressed in human Flp-In T-REx293 cells by Fura2-AM dye based calcium mobilization assay
Measurement: EC50 = 380.19 nM (pChEMBL 6.420000076293945)
Descriptors: MW 355.39, ALogP 4.33, HBA 3, HBD 0, PSA 46.61, QED 0.7
Compound (SMILES): N#CCC1(n2ccc(-c3ncnc4[nH]ccc34)c2)CN(C2CCN(C(=O)c3cccs3)CC2)C1
InChIKey: MKLXXIKQPHGDKB-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: In Vitro JAK Kinase Assay: Compounds herein were tested for inhibitory activity of JAK targets according to the following in vitro assay described in Park et al., Analytical Biochemistry 1999, 269, 94-104. The catalytic domains of human JAK1 (a.a. 837-1142), Jak2 (a.a. 828-1132) and Jak3 (a.a. 781-1124) with an N-terminal His tag were expressed using baculovirus in insect cells and purified. The catalytic activity of JAK1, JAK2 or JAK3 was assayed by measuring the phosphorylation of a biotinylated peptide. The phosphorylated peptide was detected by homogenous time resolved fluorescence (HTRF). IC50s of compounds were measured for each kinase in the 40 microL reactions that contain the enzyme, ATP and 500 nM peptide in 50 mM Tris (pH 7.8) buffer with 100 mM NaCl, 5 mM DTT, and 0.1 mg/mL (0.01%) BSA. For the 1 mM IC50 measurements, ATP concentration in the reactions was 1 mM. Reactions were carried out at room temperature for 1 hr and then stopped with 20 μL 45 mM EDTA, 300 nM SA-APC, 6 nM Eu-Py20 in assay buffer (Perkin Elmer, Boston, Mass.). Binding to the Europium labeled antibody took place for 40 minutes and HTRF signal was measured on a Fusion plate reader (Perkin Elmer, Boston, Mass.).
Measurement: IC50 = 12.5 nM (pChEMBL 7.900000095367432)
Descriptors: MW 471.59, ALogP 3.72, HBA 7, HBD 1, PSA 93.84, QED 0.48
Compound (SMILES): O=C(Cn1cc(NC(=O)c2cnn3cccnc23)c(-c2cc(Cl)ccc2OC(F)F)n1)NCC1(CO)CC1
InChIKey: UWAUFJMRLQWZHG-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: JAK Enzyme Assay: The activity of the isolated recombinant JAK1 and JAK2 kinase domain was measured by monitoring phosphorylation of a peptide derived from JAK3 (Val-Ala-Leu-Val-Asp-Gly-Tyr-Phe-Arg-Leu-Thr-Thr, fluorescently labeled on the N-terminus with 5-carboxyfluorescein) using the Caliper LabChip® technology (Caliper Life Sciences, Hopkinton, Mass.). To determine inhibition constants (Ki), compounds were diluted serially in DMSO and added to 50 μL kinase reactions containing purified enzyme (1.5 nM JAK1, or 0.2 nM JAK2), 100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 1.5 μM peptide substrate, ATP (25 μM), 10 mM MgCl2, 4 mM DTT at a final DMSO concentration of 2%. Reactions were incubated at 22° C. in 384-well polypropylene microtiter plates for 30 minutes and then stopped by addition of 25 μL of an EDTA containing solution (100 mM HEPES buffer (pH 7.2), 0.015% Brij-35, 150 mM EDTA), resulting in a final EDTA concentration of 50 mM. After termination of the kinase reaction, the proportion of phosphorylated product was determined as a fraction of total peptide substrate using the Caliper LabChip® 3000 according to the manufacturer's specifications.
Measurement: Ki = 0.55 nM (pChEMBL 9.260000228881836)
Descriptors: MW 545.93, ALogP 2.99, HBA 9, HBD 3, PSA 135.67, QED 0.28
Compound (SMILES): CO[C@H]1CC[C@H](CC(=O)N[C@H]2CC[C@H](CCN3CCC(c4noc5cc(Cl)ccc45)CC3)CC2)C1
InChIKey: FMGBBVCKDCTAKW-TZYAJKAJSA-N
Target: D(3) dopamine receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35462
Assay: Radioligand Binding Assay: Aliquots of membrane preparations were thawed at RT, resuspended in assay buffer (D2, D3: 50 mM Tris-HCl, 120 mM NaCl, 5 mM MgCl2, 1 mM EDTA, 5 mM KCl, 1.5 mM CaCl2, pH=7.4; 5-HT2A: 50 mM Tris-HCl, 10 mM MgCl2, 1 mM EGTA, pH=7.4), homogenized with a Polytron for 20-30 sec at 12,000 rpm and adjusted to a final concentration of approximately 7.5 ug protein/well (D2, D3) and 15 ug protein/well (5-HT2A), respectively.The binding affinity (Ki) of the compounds was determined using radioligand binding. Membranes were incubated in a total volume of 200 ul with a fixed concentration of radioligand (final concentration approximately 0.7 nM [3H]-spiperone for D2, 0.5 nM [3H]-spiperone for D3, and 1.1 nM [3H]-ketanserin for 5-HT2A) and ten concentrations of test compound in ranging between 10 uM-0.1 nM for 1 h at RT. At the end of the incubation, the reaction mixtures were filtered on to unifilter 96-well white microplates with bonded GF/C filters.
Measurement: Ki = 10 nM (pChEMBL 8)
Descriptors: MW 502.1, ALogP 5.93, HBA 5, HBD 1, PSA 67.6, QED 0.49
Compound (SMILES): CN(C)C(=O)c1ccccc1OCC(=O)NCC(O)CN1CCc2ccccc2C1
InChIKey: CGZXMHHOKJUGRT-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 5 (SINGLE PROTEIN), Homo sapiens
UniProt: O14744
Assay: Inhibition of PRMT5 in human Z138 cells assessed as decrease in di-methyl arginine levels after 96 hrs by Western blot method
Measurement: EC50 = 101 nM (pChEMBL 7)
Descriptors: MW 411.5, ALogP 1.3, HBA 5, HBD 2, PSA 82.11, QED 0.68
Compound (SMILES): CCN(CC)c1ccc(-c2cc(-c3ccc(C(=O)O)cc3)ccc2CCCO)cc1C(C)(C)C.Cl
InChIKey: XPDSFZHGFFIXFG-UHFFFAOYSA-N
Target: Retinoic acid receptor beta (SINGLE PROTEIN), Homo sapiens
UniProt: P10826
Assay: Activation of RARbeta (unknown origin) expressed in human HeLa cells co-expressing ERE-beta-Glob-Luc-SV-Neo incubated for 18 hrs by luciferase reporter gene assay
Measurement: Kd = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 459.63, ALogP 6.79, HBA 3, HBD 2, PSA 60.77, QED 0.37
Compound (SMILES): O=C(Nc1nc2ccccc2s1)c1cccc2c1CN(c1ccc(-c3cnn(Cc4ccccc4[N+](=O)[O-])c3)c(C(=O)O)n1)CC2
InChIKey: ODFZPPOXPFFJGT-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: FL5.12 Cellular Assay: The efficacy of the compounds of Formula (I) can also be determined in cell-based killing assays using a variety of cell lines and mouse tumor models. For example, their activity on cell viability can be assessed on a panel of cultured tumorigenic and non-tumorigenic cell lines, as well as primary mouse or human cell populations. In one exemplary set of conditions, mouse FL5.12 cells transfected with Bcl-XL were cultured under standard conditions in RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3). For assaying the compound activity, the cells were exchanged into an IL-3-depleted deprivation media, which was identical to the growth media except for the absence of FBS and WEHI-3B conditional media, for 2 days. Then the cells were exchanged to 3% FBS assay media (RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3).
Measurement: EC50 = 741 nM (pChEMBL 6.130000114440918)
Descriptors: MW 631.67, ALogP 6.02, HBA 10, HBD 2, PSA 156.38, QED 0.15
Compound (SMILES): Cc1nn(C2CCCCC2)c2sc(C(=O)NC3CCC(C(=O)N4CCOCC4)CC3)cc12
InChIKey: REAHLLQWVTXAAZ-UHFFFAOYSA-N
Target: Phosphodiesterase 4B (SINGLE PROTEIN), Mus musculus
UniProt: Q8VBU5
Assay: Inhibition Assay: The PDE 4 (phosphodiesterase IV) inhibiting effect of the compounds of the present invention was performed by the following method, which was modified assay method described in Biochemical. Pharmacol. 48(6), 1219-1223 (1994).  (1) The active fraction of PDE 4 (phosphodiesterase IV) was obtained. That is, the livers obtained from three Balb/c mice (male, 12 weeks: obtainable from CLEA Japan, Inc.) were suspended with 30 mL of buffer solution B [20 mM of bis-tris, 5 mM of 2-mercaptoethnol, 2 mM of benzamidine, 2 mM of EDTA, 0.1 mM of 4-(2-aminoethyl)benzensulfonyl hydrochloride, 50 mM of sodium acetate; pH 6.5], then homogenized by Polytron homogenizer. The homogenate were centrifuged under 25,000xG for 10 minutes at 4 C. The supernatant was separated and thus obtained supernatant was further centrifuged under 100,000xG for 60 minutes at 4Â° C., and then filtrated with 0.2 um filter to obtain the soluble fraction.
Measurement: IC50 = 2700 nM (pChEMBL 5.570000171661377)
Descriptors: MW 458.63, ALogP 4.06, HBA 6, HBD 1, PSA 76.46, QED 0.75
Compound (SMILES): COc1ccc(C(=O)C2CCN(C3CCN(Cc4nc5c(cnn5C)c(=O)[nH]4)C3=O)CC2)cc1
InChIKey: TYQINFQXCFVKCF-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase tankyrase-1 (SINGLE PROTEIN), Homo sapiens
UniProt: O95271
Assay: Biochemical Assay: The autoparsylation activity of the TNKS 1/2 or PARP1/2 enzymes was measured by the liquid chromatography-mass spectrometry (LC/MS) detection of nicotinamide as readout. Compound activity in inhibiting the TNKS and PARP autoparsylation was evaluated by IC50 measurements. In the compound screening assays, the reaction is composed of 5 uL of compound in 8-point serial dilutions with concentrations ranging from 0.0086 to 18.75 uM, 20 nM of purified enzyme, and 250 uM of p-NAD+ in the 1x Assay Buffer. After 60 min incubation at room temperature, the reactions were quenched by the addition of 10 uL of 5x quenching solution (20% formic acid and 500 nM [d]-nicotinamide in water). For the background control wells, 10 uL of the 5x quenching solution per well was added prior to the addition of p-NAD+. The % Inhibition was calculated as: (Control-Sample)/(Control-Background)*100. "Control" is the average value of 8 wells without compound; and "Background" is the average of 8 wells mixed with 5x quenching solution measured prior to initiation of the reaction.
Measurement: IC50 = 22 nM (pChEMBL 7.659999847412109)
Descriptors: MW 464.53, ALogP 1.36, HBA 8, HBD 1, PSA 113.42, QED 0.55
Compound (SMILES): COc1cc(C(=O)N2CC[C@H]2c2nc(-c3cccc(Cl)c3C)no2)c(-n2nccn2)cc1Cl
InChIKey: TWOGTHWXONLTPL-KRWDZBQOSA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: Intracellular Calcium Assay: Chinese hamster ovary (CHO) cells expressing the human orexin-1 receptor and the human orexin-2 receptor, respectively, are grown in culture medium (Ham F-12 with L-Glutamine) containing 300 μg/mL G418, 100 U/mL penicillin, 100 μg/mL streptomycin and 10% heat inactivated fetal calf serum (FCS). The cells are seeded at 20,000 cells/well into 384-well black clear bottom sterile plates (Greiner). The seeded plates are incubated overnight at 37° C. in 5% CO2. Human orexin-A as an agonist is prepared as 1 mM stock solution in MeOH:water (1:1), diluted in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/L and 20 mM HEPES for use in the assay at a final concentration of 3 nM. Antagonists are prepared as 10 mM stock solution in DMSO, then diluted in 384-well plates using DMSO followed by a transfer of the dilutions into in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/L and 20 mM HEPES. On the day of the assay, 50 μL of staining buffer (HBSS containing 1% FCS, 20 mM HEPES, NaHCO3: 0.375 g/L, 5 mM probenecid (Sigma) and 3 μM of the fluorescent calcium indicator fluo-4 AM (1 mM stock solution in DMSO, containing 10% pluronic) is added to each well. The 384-well cell-plates are incubated for 50 min at 37° C. in 5% CO2 followed by equilibration at rt for 30 min before measurement. Within the Fluorescent Imaging Plate Reader (FLIPR Tetra, Molecular Devices), antagonists are added to the plate in a volume of 10 μL/well, incubated for 120 min and finally 10 μL/well of agonist is added. Fluorescence is measured for each well at 1 second intervals, and the height of each fluorescence peak is compared to the height of the fluorescence peak induced by 3 nM orexin-A with vehicle in place of antagonist.
Measurement: IC50 = 6 nM (pChEMBL 8.220000267028809)
Descriptors: MW 485.33, ALogP 4.53, HBA 8, HBD 0, PSA 99.17, QED 0.41
Compound (SMILES): CN1C(=N)N[C@](C)(c2sc(-c3ccnc(-c4ccncc4)c3)cc2F)CS1(=O)=O
InChIKey: NRJDTUXBLSXRTB-IBGZPJMESA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: HTRF FRET Assay: Inhibitor compounds, prepared at 3× the desired final concentration in 1×BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO are pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1×BACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30° C. The assay is initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 μM for 4 μM for autoBACE-2) prepared in 1×BACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C. DMSO is present at 5% final concentration in the assay. Following laser excitation of sample wells at 320 nm, the fluorescence signal at 620 nm is collected for 400 ms following a 50 μs delay on a RUBYstar HTRF plate reader (BMG Labtechnologies).
Measurement: Ki = 123 nM (pChEMBL 6.909999847412109)
Descriptors: MW 431.52, ALogP 3.03, HBA 6, HBD 2, PSA 99.04, QED 0.66
Compound (SMILES): c1ccc(C2CC2NC2CCNCC2)cc1
InChIKey: BASFYRLYJAZPPL-UHFFFAOYSA-N
Target: CDGSH iron-sulfur domain-containing protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NZ45
Assay: Displacement of [3H]rosiglitazone from recombinant human C-terminal His-tagged MitoNEET cytosolic domain (32 to 108 residues) expressed in Escherichia coli BL21 by Cheng-Prusoff analysis
Measurement: Ki = 148 nM (pChEMBL 6.829999923706055)
Descriptors: MW 216.33, ALogP 1.88, HBA 2, HBD 2, PSA 24.06, QED 0.81
Compound (SMILES): COc1cccc(OC)c1C1CCCC(=O)N1Cc1ccnc(Br)c1
InChIKey: DSSSAICYQZFDPR-UHFFFAOYSA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: In Vitro Assay: Intracellular Calcium Measurements : In Vitro Assay: Intracellular Calcium Measurements:Chinese hamster ovary (CHO) cells expressing the human orexin-1 receptor and the human orexin-2 receptor, respectively, are grown in culture medium (Ham F-12 with L-Glutamine) containing 300 g/ml G418, 100 U/ml penicillin, 100 g/ml streptomycin and 10% heat inactivated fetal calf serum (FCS). The cells are seeded at 20,000 cells/well into 384-well black clear bottom sterile plates (Greiner). The seeded plates are incubated overnight at 37° C. in 5% CO2. Human orexin-A as an agonist is prepared as 1 mM stock solution in MeOH:water (1:1), diluted in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/l and 20 mM HEPES for use in the assay at a final concentration of 3 nM.Antagonists are prepared as 10 mM stock solution in DMSO, then diluted in 384-well plates using DMSO followed by a transfer of the dilutions into in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/l and 20 mM HEPES. On the day of the assay, 50 μl of staining buffer (HBSS containing 1% FCS, 20 mM HEPES, NaHCO3: 0.375 g/l, 5 mM probenecid (Sigma) and 3 M of the fluorescent calcium indicator fluo-4 AM (1 mM stock solution in DMSO, containing 10% pluronic) is added to each well. The 384-well cell-plates are incubated for 50 min at 37° C. in 5% CO2 followed by equilibration at RT for 30 min before measurement.
Measurement: IC50 = 1150 nM (pChEMBL 5.940000057220459)
Descriptors: MW 405.29, ALogP 4.12, HBA 4, HBD 0, PSA 51.66, QED 0.7
Compound (SMILES): CCCOc1ccc(-c2nc(-c3cccc4c3CCN4CC(N)(CO)CO)no2)cc1OCCC
InChIKey: NBOTXMJUPYPBQH-UHFFFAOYSA-N
Target: Sphingosine 1-phosphate receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99500
Assay: Calcium Mobilization Assay: Selected Compounds of the Examples were evaluated at Millipore Corporation, USA, using S1P1 receptor; [35S]-GTPgamaS binding assay. A [35S]-GTPgamaS binding assay at Millipore was conducted by GPCR Profiler Custom Service Laboratory, Temecula, Calif., Millipore, Inc. to monitor dose-dependent agonist selectivity for selected Examples against the S1P1 receptors. The assay was completed with sample compounds subjected to an eight-point, four-fold dose response curve with starting concentration of 10 uM. Selectivity was determined upon initial addition of compounds followed by a 30 minute incubation at 30° C. Following compound incubation, bounded [35S]-GTPgamaS was determined by filtration and scintillation counting. Independently, selected compounds were evaluated for S1P1 and S1P3 agonistic activity. The S1P1 assay system was GTPgama-S35 binding in membranes from CHO K1 cells, expressing S1P1 human receptor. The S1P3 assay system was calcium mobilization in CHO K1 cells expression.
Measurement: EC50 = 310 nM (pChEMBL 6.510000228881836)
Descriptors: MW 482.58, ALogP 3.03, HBA 9, HBD 3, PSA 127.1, QED 0.36
Compound (SMILES): COc1cc(OC)c(C)c(N(CCCN2CCCC2)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1.Cl
InChIKey: OKFNCOXKQXFNDZ-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 = 6918.31 nM (pChEMBL 5.159999847412109)
Descriptors: MW 486.62, ALogP 4.98, HBA 8, HBD 0, PSA 68.54, QED 0.33
Compound (SMILES): COc1ccc2cc(CCc3c(C(=O)O)[nH]c(=O)[nH]c3=O)c(OC)cc2c1.NCCO
InChIKey: BHTFTQHUPFNVLR-UHFFFAOYSA-N
Target: Dihydroorotate dehydrogenase (quinone), mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: Q02127
Assay: Inhibition of human dihydroorotate dehydrogenase assessed as reduction in orotate production using L-dihydroorotate as substrate measured after 20 mins by spectrophotometric method
Measurement: IC50 = 47.8 nM (pChEMBL 7.320000171661377)
Descriptors: MW 370.36, ALogP 1.72, HBA 5, HBD 3, PSA 121.48, QED 0.61
Compound (SMILES): COc1cc(OC)cc(N(CC(C)NC(C)C)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1.Cl
InChIKey: WQPXPTDVBCLUDQ-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Inhibition of FGFR3 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 460.58, ALogP 4.57, HBA 8, HBD 1, PSA 77.33, QED 0.39
Compound (SMILES): CC([C@H]1CC[C@H](C)CC1)n1c(N2C[C@@H](O)C[C@@H]2C)nc2cc(-c3nc(=O)o[nH]3)nc(-c3cncc(Cl)c3)c21
InChIKey: NNJDDNCVQBYTOS-WLNPMRSCSA-N
Target: E3 ubiquitin-protein ligase Mdm2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00987
Assay: FRET Assay: Methods: An HDM2 FRET assay was developed to assess the compounds' inhibitory activity towards binding of p53 protein. A truncated version of HDM2 with residues 17 to 125 (containing p53 binding surface, Science (1994) 265, 346-355), with N-terminal His and Thioredoxin tag was generated in pET32a expression vector and expressed in E. coli strain BL21(DE3)Rosetta. Protein was purified using Ni-affinity chromatography, followed by size exclusion chromatography using Superdex 75 26/60 column. To assess inhibition of p53 binding to HDM2, a FITC labeled 8-mer peptide (SEQ ID NO:1: Ac-Phe-Arg-Dpr-Ac6c-(6-Br)Trp-Glu-Glu-Leu-NH2; Anal Biochem. 2004 Aug. 1; 331(1):138-46) with strong affinity towards the p53 binding pocket of HDM2 was used. The HDM2 assay buffer contained 1x Phosphate Buffered Saline (Invitrogen, Cat#14190), 0.01% BSA (Jackson ImmunoResearch, Cat#001-000-162), 0.01% Tween-20. In the 1x assay buffer recombinant HDM2 protein, peptide and Lumi-4-Tb Cryptate-conjugate mouse.
Measurement: IC50 = 3 nM (pChEMBL 8.520000457763672)
Descriptors: MW 538.05, ALogP 4.84, HBA 9, HBD 2, PSA 125.96, QED 0.37
Compound (SMILES): COc1cc(-c2cnc(N)c(-c3cn(C(C)C)nn3)c2)ccc1C(=O)N1CCOCC1
InChIKey: WTMSPLKBPPJPHJ-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: Kinase Activity Assay: The kinase reaction buffer consists of 50 mM Tris pH 8.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA, and 2 mM DTT. In the assay, 1 nM LRRK2 WT or 250 pM LRRK2 G2019S kinase is incubated with the test compound (typically at 0 to 30 μM) for 30 minutes before the kinase reaction is initiated by addition of 1.3 mM ATP and 0.4 μM fluorescein-LRRKtide. The reaction mixture (20 μl total volume) is incubated for 2 hours at 30 °C. before the reaction is terminated by addition of 10 mM EDTA and 1 nM terbium-labelled anti-phospho-LRRKtide antibody (final volume 20 μl). The mixture is further incubated for 30 minutes at RT. TR-FRET is measured by excitation of the terbium-donor with 340 nm light and subsequent (delay time 100 μs) measurement of terbium and fluorescein emission at 495 nm and 520 nm, respectively, over a time window of 1000 μs. The measurement is repeated 10 times for fluorescein and 10 times for terbium emission with a 2000 μs time window between repeats. TR-FRET measurements are performed on a Biomek Synergy plate. The TR-FRET signal is calculated as the emission-ratio at 520 nm over 495 nm.
Measurement: IC50 = 91 nM (pChEMBL 7.039999961853027)
Descriptors: MW 422.49, ALogP 2.65, HBA 8, HBD 1, PSA 108.39, QED 0.67
Compound (SMILES): Cc1c(-c2ccc(N3CCc4cccc(C(=O)Nc5nc6ccccc6s5)c4C3)nc2C(=O)O)cnn1CC1(OCCS(C)(=O)=O)CCCC(C)(C)C1
InChIKey: YAPPRYLJJFBFDI-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: FL5.12 Cellular Assay: The efficacy of the compounds of Formula (I) can also be determined in cell-based killing assays using a variety of cell lines and mouse tumor models. For example, their activity on cell viability can be assessed on a panel of cultured tumorigenic and non-tumorigenic cell lines, as well as primary mouse or human cell populations. In one exemplary set of conditions, mouse FL5.12 cells transfected with Bcl-XL were cultured under standard conditions in RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3). For assaying the compound activity, the cells were exchanged into an IL-3-depleted deprivation media, which was identical to the growth media except for the absence of FBS and WEHI-3B conditional media, for 2 days. Then the cells were exchanged to 3% FBS assay media (RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3).
Measurement: EC50 = 0.2 nM (pChEMBL 9.699999809265137)
Descriptors: MW 756.95, ALogP 6.78, HBA 11, HBD 2, PSA 156.61, QED 0.15
Compound (SMILES): Cc1noc(C)c1-c1cc(C(O)(c2nccs2)c2nccs2)c2nc(C3CC3)[nH]c2c1
InChIKey: TZQMYCBSCKPPKG-UHFFFAOYSA-N
Target: Bromodomain-containing protein 4 (SINGLE PROTEIN), Homo sapiens
UniProt: O60885
Assay: Ligand KI Assay: The labeled ligand specifically binds BRD4-1 and BRD4-2 and can be displaced by a small molecule inhibitor that shares a similar or overlapping binding site. BRD4-I and BRD4-2 were expressed and purified from Escherichia coli as N-terminal His6, tagged proteins. A Eu-cryptate labeled anti-His antibody (Perkin Elmer) was used to specifically bind BRD4. Binding of BRD4 to the labeled probe/ligand resulted in an increase in FRET signal whereas displacement of this labeled ligand from BRD4 with a small molecule inhibitor resulted in a decrease in FRET signal. Assays were performed in 50 mM Hepes (pH 7.5), 150 mM NaCl, 0.1 mg/ml BSA, 0.01% (v/v) Brij, 0.5% (v/v) DMSO and 10 nM labeled ligand at the following concentrations for each BRD4 isoform: 2 nM BRD4-1 and 0.5 nM BRD4-2. After an assay reaction time of 60 minutes at 25° C., binding was measured with 2 nM Eu-cryptate labeled anti-His antibody. TR-FRET signal was detected on an Envision plate reader (Ex: 320 nm; Em: 615/665 nm; 100 us delay and 200 us read window). Data were normalized based on a positive (2 uM I-BET) and negative (DMSO) controls and IC50 values were calculated from the fit of the dose-response curves to a four-parameter equation. All IC50 values represent geometric mean values of a minimum of four determinations. The IC50 values were converted to Ki values (dissociation constant for BRD4-inhibitor complex) using the Cheng and Prusoff equation for a competitive inhibitor mode of action.
Measurement: IC50 = 12.4 nM (pChEMBL 7.909999847412109)
Descriptors: MW 449.56, ALogP 4.91, HBA 8, HBD 2, PSA 100.72, QED 0.4
Compound (SMILES): COC1(Cn2ncc(-c3ccc(N4CCc5cccc(C(=O)Nc6nc7ccccc7s6)c5C4)nc3C(=O)O)c2C)CCCC(C)(C)C1
InChIKey: PCJOKOQMKUYYRS-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: FL5.12 Cellular Assay: The efficacy of the compounds of Formula (I) can also be determined in cell-based killing assays using a variety of cell lines and mouse tumor models. For example, their activity on cell viability can be assessed on a panel of cultured tumorigenic and non-tumorigenic cell lines, as well as primary mouse or human cell populations. In one exemplary set of conditions, mouse FL5.12 cells transfected with Bcl-XL were cultured under standard conditions in RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3). For assaying the compound activity, the cells were exchanged into an IL-3-depleted deprivation media, which was identical to the growth media except for the absence of FBS and WEHI-3B conditional media, for 2 days. Then the cells were exchanged to 3% FBS assay media (RPMI with 2 mM glutamine, 1% 100 mM sodium pyruvate, 2% 1 M HEPES, 4 uL/L of beta;-mercaptoethanol, 1% penicillin-streptomycin, 10% FBS, and 10% WEHI-3B conditioned media (for IL-3).
Measurement: EC50 = 0.3 nM (pChEMBL 9.520000457763672)
Descriptors: MW 664.83, ALogP 7.36, HBA 9, HBD 2, PSA 122.47, QED 0.18
Compound (SMILES): O=C(CCCCCCCOc1ccc2cc1COC/C=C/COCc1cccc(c1)-c1ccnc(n1)N2)NO
InChIKey: MCVHEQSOIJARSD-VOTSOKGWSA-N
Target: Protein deacetylase HDAC6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UBN7
Assay: Inhibition of full length human recombinant N-terminal GST-tagged HDAC6 (1 to 1215 residues) expressed in sf9 cells preincubated with enzyme followed by fluorogenic Arg-His-Lys-Lys(Ac)-AMC substrate addition measured after 2 hrs by fluorescence assay
Measurement: IC50 = 15.8 nM (pChEMBL 7.800000190734863)
Descriptors: MW 532.64, ALogP 5.72, HBA 8, HBD 3, PSA 114.83, QED 0.14
Compound (SMILES): O=c1[nH]c(C2CCC2c2ccccn2)nc2c1cnn2C1CCOCC1
InChIKey: IVFWFHHENYUJLU-UHFFFAOYSA-N
Target: High affinity cGMP-specific 3',5'-cyclic phosphodiesterase 9A (SINGLE PROTEIN), Homo sapiens
UniProt: O76083
Assay: Scintillation Proximity Assay: The PDE9A2 enzymatic activity assay was run as scintillation proximity assay (SPA), in general according to the protocol of the manufacturer (GE Healthcare, former Amersham Biosciences, product number: TRKQ 7100). As enzyme source, lysate (PBS with 1% Triton X-100 supplemented with protease inhibitors, cell debris removed by centrifugation at 13.000 rpm for 30 min) of SF 9 cell expressing the human PDE9A2 was used. The total protein amount included in the assay varied upon infection and production efficacy of the SF9 cells and lay in the range of 0.1-100 ng. In general, the assay conditions were as follows: total assay volume: 40 microlitre; protein amount: 0.1-50 ng; substrate concentration (cGMP): 20 nanomolar; ~1 mCi/l; incubation time: 60 min at room temperature; final DMSO concentration: 0.2-1%. The assays were run in 384-well format. The test reagents as well as the enzyme and the substrate were diluted in assay buffer. The assay buffer contained 50 mM Tris, 8.3 mM MgCl2, 1.7 mM EGTA, 0.1% BSA, 0.05% Tween 20; the pH of assay buffer was adjusted to 7.5. The reaction was stopped by applying a PDE9 specific inhibitor (e.g. compounds according to WO 2004/099210 or WO 2004/099211, like one of the enantiomers of example 37, e.g. 1-(2-Chlorophenyl)-6-[2R)-3,3,3-trifluoro-2-methyl-propyl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one) in excess.
Measurement: IC50 = 7 nM (pChEMBL 8.149999618530273)
Descriptors: MW 351.41, ALogP 2.53, HBA 6, HBD 1, PSA 85.69, QED 0.78
Compound (SMILES): CCc1c(N)ncnc1N1CCC(c2nc(-c3ccc(Cl)cc3)cn2CCN2CCCC2)CC1
InChIKey: PZOKYBNWYFILSM-UHFFFAOYSA-N
Target: RAC-alpha serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P31749
Assay: AKT Enzyme Assay: A TTP Mosquito liquid handling instrument is used to place 125 nl of the appropriate concentration of inhibitor in 100% DMSO (for a dose response curve calculation) into each well of a 384-well plate. To this reaction components are added to a final volume of 12.5 .mu.l: 0.1 ng/.mu.l His-AKT (Full Length), (Invitrogen, Part # P2999, Lot #641228C). 160 uM ATP (Fluka, 02055) 1 mM DTT (Sigma, D0632) 1 mM MgCl2 (Sigma, M1028) 1 .mu.M substrate peptide (sequence FITC-AHA-GRPRTSSFAEG-NH2), synthesized by Tufts Peptide Synthesis service. 100 mM HEPES pH 7.5 (Calbiochem, 391338)0.015% Brij-35 (Sigma, B4184)The reaction is incubated for 90 min at 25 C, and then stopped by the addition of 70 .mu.l of Stop buffer (100 mM HEPES pH 7.5, 0.015% Brij-35, 10 mM EDTA (Sigma, E7889)).The plate is read on a Caliper LC 3000 in an Off-Chip mobility shift assay format, using the following parameters for a 12-sipper chip: screening pressure -2.3 psi, upstream voltage -500, and downstream voltage -3000.
Measurement: IC50 = 190 nM (pChEMBL 6.71999979019165)
Descriptors: MW 480.06, ALogP 4.62, HBA 7, HBD 1, PSA 76.1, QED 0.53
Compound (SMILES): O=C(NC1(CO)CC1)c1cc(OCC2CC2)c(N2CC(F)(F)C2)cn1
InChIKey: WOXPSIFPMYUHQL-UHFFFAOYSA-N
Target: Cannabinoid receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21554
Assay: cAMP Assay: CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hours prior to the experiment 50.000 cells per well in a black 96 well plate with flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No. 31331), 1×HT supplement, with 10% fetal calf serum and incubated at 5% CO2 and 37° C. in a humidified incubator. The growth medium was exchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX and incubated at 30° C. for 30 min. Compounds were added to a final assay volume of 100 μl and incubated for 30 min at 30° C. Using the cAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped by the addition of 50 μl lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5% NP40, 10% NaN3) and 50 μl detection solutions (20 μM mAb Alexa700-cAMP 1:1, and 48 μM Ruthenium-2-AHA-cAMP) and shaken for 2 h at room temperature. The time-resolved energy transfer is measured by a TRF reader (Evotec Technologies GmbH), equipped with a ND:YAG laser as excitation source. The plate is measured twice with the excitation at 355 nm and at the emission with a delay of 100 ns and a gate of 100 ns, total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth 75 nm), respectively. The FRET signal is calculated as follows: FRET=T730-Alexa730-P(T645-B645) with P=Ru730-B730/Ru645-B645, where T730 is the test well measured at 730 nM, T645 is the test well measured at 645 nm, B730 and B645 are the buffer controls at 730 nm and 645 nm, respectively. cAMP content is determined from the function of a standard curve spanning from 10 μM to 0.13 nM cAMP.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 353.37, ALogP 1.58, HBA 5, HBD 2, PSA 74.69, QED 0.78
Compound (SMILES): O=C(Nc1ccc(OC(F)(F)F)cc1)c1cnc(N2CCNC3(CC3)C2)c(-c2ccn[nH]2)c1
InChIKey: GIZTWOXMTDJYBI-UHFFFAOYSA-N
Target: Tyrosine-protein kinase ABL1 (SINGLE PROTEIN), Homo sapiens
UniProt: P00519
Assay: Caliper Assay: The assay plates were prepared by addition of 50 nL per well of compound solution in 90% DMSO. The kinase reactions were started by stepwise addition of 4.5 μL per well of peptide/ATP-solution (50 mM HEPES, pH 7.5, 1 mM DTT, 0.02% BSA, 0.6% DMSO, 10 mM beta-glycerophosphate, and 10 μM sodium orthovanadate, 20 mM MgCl2, 2 mM MnCl2, 4 μM ATP, 4 μM peptide (FITC-Ahx-EAIYAAPFAKKK-NH2)) and 4.5 μL per well of enzyme solution (50 mM HEPES, pH 7.5, 1 mM DTT, 0.02% BSA, 0.6% DMSO, 10 mM beta-glycerophosphate, and 10 μM sodium orthovanadate, 20 mM MgCl2, 2 mM MnCl2, 3.5 nM ABL (ABL(64-515), produced in-house from E. coli)). Kinase reactions were incubated at 30° C. for 60 minutes and subsequently terminated by addition of 16 μL per well of stop solution (100 mM HEPES pH 7.5, 5% DMSO, 0.1% Caliper coating reagent, 10 mM EDTA, and 0.015% Brij35). Plates with terminated kinase reactions were transferred to the Caliper LC3000 workstations for reading. Phosphorylated and unphosphorylated peptides were separated using the Caliper microfluidic mobility shift technology. Briefly, samples from terminated kinase reactions were applied to the chip. Analytes are transported through the chip by constant buffer flow and the migration of the substrate peptide is monitored by the fluorescence signal of its label. Phosphorylated peptide (product) and unphosphorylated peptide (substrate) are separated in an electric field by their charge/mass ratio.
Measurement: IC50 = 8.3 nM (pChEMBL 8.079999923706055)
Descriptors: MW 458.44, ALogP 3.56, HBA 6, HBD 3, PSA 95.17, QED 0.54
Compound (SMILES): COc1cccc(-c2noc([C@@H]3CCN3C(=O)c3cc(C)ccc3-n3nccn3)n2)c1
InChIKey: DREBVPAPAKBWSB-IBGZPJMESA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: Intracellular Calcium Assay: Chinese hamster ovary (CHO) cells expressing the human orexin-1 receptor and the human orexin-2 receptor, respectively, are grown in culture medium (Ham F-12 with L-Glutamine) containing 300 μg/mL G418, 100 U/mL penicillin, 100 μg/mL streptomycin and 10% heat inactivated fetal calf serum (FCS). The cells are seeded at 20,000 cells/well into 384-well black clear bottom sterile plates (Greiner). The seeded plates are incubated overnight at 37° C. in 5% CO2. Human orexin-A as an agonist is prepared as 1 mM stock solution in MeOH:water (1:1), diluted in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/L and 20 mM HEPES for use in the assay at a final concentration of 3 nM. Antagonists are prepared as 10 mM stock solution in DMSO, then diluted in 384-well plates using DMSO followed by a transfer of the dilutions into in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/L and 20 mM HEPES. On the day of the assay, 50 μL of staining buffer (HBSS containing 1% FCS, 20 mM HEPES, NaHCO3: 0.375 g/L, 5 mM probenecid (Sigma) and 3 μM of the fluorescent calcium indicator fluo-4 AM (1 mM stock solution in DMSO, containing 10% pluronic) is added to each well. The 384-well cell-plates are incubated for 50 min at 37° C. in 5% CO2 followed by equilibration at rt for 30 min before measurement. Within the Fluorescent Imaging Plate Reader (FLIPR Tetra, Molecular Devices), antagonists are added to the plate in a volume of 10 μL/well, incubated for 120 min and finally 10 μL/well of agonist is added. Fluorescence is measured for each well at 1 second intervals, and the height of each fluorescence peak is compared to the height of the fluorescence peak induced by 3 nM orexin-A with vehicle in place of antagonist.
Measurement: IC50 = 25 nM (pChEMBL 7.599999904632568)
Descriptors: MW 416.44, ALogP 3.22, HBA 8, HBD 0, PSA 99.17, QED 0.49
Compound (SMILES): CC(C(=O)Nc1ccc(C(=O)O)cc1)n1cc(F)c(-c2cc(Cl)ccc2C#N)cc1=O
InChIKey: BFAOKCDIFBZLTD-UHFFFAOYSA-N
Target: Plasma kallikrein (SINGLE PROTEIN), Homo sapiens
UniProt: P03952
Assay: Inhibition Assay: Test substances are dissolved in dimethyl sulphoxide and serially diluted in dimethyl sulphoxide (3000 μM to 0.0078 μM; resulting final concentrations in the test: 50 μM to 0.00013 μM). In each case 1 μl of the diluted substance solutions are placed into the wells of white microtitre plates from Greiner (384 wells). 20 μl of assay buffer (50 mM Tris/HCl pH 7.4; 100 mM sodium chloride solution; 5 mM of calcium chloride solution; 0.1% of bovine serum albumin) and 20 μl of plasma kallikrein from Kordia (0.6 nM in assay buffer) are then added successively. After 15 min of incubation, the enzyme reaction is started by addition of 20 μl of the substrate H-Pro-Phe-Arg-AMC dissolved in assay buffer (10 μl in assay buffer) from Bachem, the mixture is incubated at room temperature (22° C.) for 30 min and fluorescence is then measured (excitation: 360 nm, emission: 460 nm).
Measurement: IC50 = 310 nM (pChEMBL 6.510000228881836)
Descriptors: MW 439.83, ALogP 4.08, HBA 5, HBD 2, PSA 112.19, QED 0.62
Compound (SMILES): OC(Cn1cnnn1)(c1ccc(F)cc1F)C(F)(F)c1ccc(-c2ccc(F)nc2)cn1
InChIKey: BPJZEQQYTJUNIW-UHFFFAOYSA-N
Target: Cytochrome P450 2C19 (SINGLE PROTEIN), Homo sapiens
UniProt: P33261
Assay: Inhibition Assay: Solutions of each test compound were separately prepared at concentrations of 20000, 6000, 2000, 600, 200, and 60 uM by serial dilution with DMSO:MeCN (50:50 v/v). The individual test compound solutions were then diluted 20-fold with DMSO:MeCN:deionized water (5:5:180 v/v/v) to concentrations of 1000, 300, 100, 30, 10, and 3 Î¼M. Mixtures of isozyme inhibitors (sulfaphenazole, tranylcypromine, and ketoconazole as specific inhibitors of isozymes 2C9, 2C19, and 3A4, respectively) were prepared containing each inhibitor at concentrations of 6000, 2000, 600, 200, 60, 20, 6, and 2 uM by serial dilution with DMSO:CH3CN (50:50 v/v). The mixed inhibitor solutions were then diluted 20-fold with DMSO: CH3CN:deionized water (5:5:180 v/v/v) to concentrations of 300, 100, 30, 10, 3, 1, 0.3, and 0.1 uM. The percent of organic solvent attributable to the test compound or inhibitor mixture in the final reaction mixture was 2% v/v. Pooled human liver microsome suspension (20 mg/mL) was diluted with phosphate buffer to obtain a 5 mg/mL suspension.
Measurement: IC50 = 61000 nM (pChEMBL 4.210000038146973)
Descriptors: MW 448.36, ALogP 3.23, HBA 7, HBD 1, PSA 89.61, QED 0.36
Compound (SMILES): COC(=O)Nc1nc2ccc(C3(O)c4ccccc4C(=O)N3Cc3cccc(Br)c3)cc2[nH]1
InChIKey: MTQYHLUEBBENHR-UHFFFAOYSA-N
Target: RAF proto-oncogene serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P04049
Assay: Inhibition of c-Raf1 (unknown origin) pre-incubated for 30 mins by luciferase-luciferin-coupled chemiluminescence assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 507.34, ALogP 4.35, HBA 5, HBD 3, PSA 107.55, QED 0.38
Compound (SMILES): COC(=O)CCC(=O)Nc1ccc2nc(NC(=O)c3cc(Cl)c(Cl)[nH]3)sc2c1
InChIKey: SUKOIIJJKMQGFR-UHFFFAOYSA-N
Target: DNA gyrase subunit B (SINGLE PROTEIN), Escherichia coli
UniProt: P0AES6
Assay: Inhibition of Escherichia coli DNA gyrase B ATP binding site assessed as reduction in supercoiling of relaxed pNO1 DNA after 30 mins in presence of biotinylated oligonucleotide by sybrGOLD staining based fluorescence assay
Measurement: IC50 = 35000 nM (pChEMBL 4.460000038146973)
Descriptors: MW 441.3, ALogP 4.08, HBA 6, HBD 3, PSA 113.18, QED 0.5
Compound (SMILES): COc1c(Nc2ncc(Cl)c(N[C@H]3[C@@H](C(N)=O)[C@@H]4C=C[C@H]3C4)n2)ccc2c1CCC[C@H](N1CCOCC1)C2
InChIKey: LIBCMUMCEYECFM-CVOKMOJFSA-N
Target: Insulin receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P06213
Assay: Inhibition of recombinant human INSR expressed in baculovirus using ATP as substrate after 15 mins by TRF assay
Measurement: IC50 = 994 nM (pChEMBL 6)
Descriptors: MW 539.08, ALogP 3.55, HBA 8, HBD 3, PSA 114.63, QED 0.36
Compound (SMILES): Cc1ccc(NC(=O)c2cccc(C(F)(F)F)c2)cc1-c1cn(C)c(=O)c(N2CCOCC2)n1
InChIKey: VUCDTBXJRRGGMN-UHFFFAOYSA-N
Target: RAF proto-oncogene serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P04049
Assay: In Vitro Enzyme Assay: Final assay conditions were 4 pM cRaf, 3 uM ATP, 10 nM biotin tagged MEK1 kinase dead protein substrate. Reactions were performed in Greiner384 well plates, catalog #784075. Reaction buffer was 50 mM Tris, pH 7.5, 50 mM NaCl, 10 mM MgCl2, 0.01% Tween-20, 0.05% BSA and 1 mM DTT. 5 ul of 2×cRaf plus MEK1 were dispensed into plates containing 0.25 ul of compound in 100% DMSO and incubated at room temperature for 30 minutes. Reactions were started by addition of 5 ul of 2×ATP. Reactions were run for two hours and terminated by addition of 10 ul of 2× antibody and Alphascreen bead mixture in Stop Buffer. Stop Buffer was 50 mM Tris, pH 7.5, 0.005% Tween-20, 25 mM EDTA.
Measurement: IC50 = 0.27 nM (pChEMBL 9.569999694824219)
Descriptors: MW 472.47, ALogP 3.86, HBA 6, HBD 1, PSA 76.46, QED 0.62
Compound (SMILES): CC1(c2cccc(Nc3nsc4cc(Br)ccc34)c2)COCC(N)=N1
InChIKey: IRNHZHZGEZSMNC-UHFFFAOYSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: BACE-2 Inhibition Assay: Recombinant BACE-2 (extracellular domain, expressed in baculovirus and purified using standard methods) at 0.1 to 1 nM concentrations is incubated with the test compound at various concentrations for 1 hour at room temperature in 100 mM acetate buffer, pH 4.5, containing 0.1% CHAPS. Activity was measured using a final concentration of 3 uM of the fluorescence-quenched substrate Q-C(HSO3)-Ile-Asp-Leu-Ala-Val-Leu-Asp-HN-CH2-CH2-Mca, where Q=2-nitro-5-amino benzoic acid and Mca=7-methoxy-4-coumarinyl acetic acid. Catalytic turnover was monitored in a Spectramax Gemini fluorescence plate reader (Molecular Devices) in black 96-well microplates using excitation/emission wavelength of 325 nm and 400 nm, respectively. Fluorescence increase was followed for 15 min, in 1 minute's intervals.
Measurement: IC50 = 1900 nM (pChEMBL 5.71999979019165)
Descriptors: MW 417.33, ALogP 4.41, HBA 6, HBD 2, PSA 72.53, QED 0.66
Compound (SMILES): COc1ccc([C@H](NC(=O)c2cc(F)c3cnc(NC(C)C(F)(F)F)nc3c2)c2cnn(C)c2)cc1F
InChIKey: ZNTSMVKYLVMTDD-FHQWGVRCSA-N
Target: Mitogen-activated protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P28482
Assay: Enzymatic Assay: Compounds were tested in an enzymatic assay using human ERK-2 (Mitogen Activated Kinase 1), recombinantly expressed as an N-terminal 6-His fusion protein in E. coli and corresponding to aa 8-360. The substrate used was the fluorescent Omnia peptide S/T17 (Invitrogen of Carlsbad, Calif.; Cat. KNZ1171C). Test compounds were diluted in DMSO in 3-fold serial dilutions at 100× final concentrations. In addition to compound, the assay contained 50 mM HEPES [pH 7.3], 10 mM MgCl2, 1 mM DTT, 0.005% Triton-X100, 5 nM ERK-2 enzyme, 6.25 μM S/T17 peptide substrate and 25 μM ATP (corresponding to the observed Km) for a total reaction volume of 25 μL. The assay was run at ambient temperature in a white 384-well polypropylene plate (Nunc, Inc of Naperville, Ill.; Cat. 267462) collecting data every 50 seconds for approximately 30 minutes on an Envision plate reader (PerkinElmer, Inc. of Waltham, Mass.); Excitation 340 nm/Emission 495 nm.
Measurement: IC50 = 13.6 nM (pChEMBL 7.869999885559082)
Descriptors: MW 520.46, ALogP 4.53, HBA 7, HBD 2, PSA 93.96, QED 0.35
Compound (SMILES): CN1C(=O)CCc2cc(-c3cncc4c3CCCC4=O)ccc21
InChIKey: DQSZXHUFCGFADZ-UHFFFAOYSA-N
Target: Cytochrome P450 11B1, mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: P15538
Assay: Cellular Enzyme Assay: The expression plasmids contains the ORF for either human/cyno CYP11B1 or CYP11B2 under the control of a suitable promoter (CMV-promoter) and a suitable resistance marker (neomycin). Using standard techniques the expression plasmid is transfected into G-402 cells and these cells are then selected for expressing the given resistance markers. Individual cell-clones are then selected and assessed for displaying the desired enzymatic activity using 11-Deoxycorticosterone (Cyp11B2) or 11-Deoxycortisol (Cyp11B1) as a substrate. G-402 cells expressing CYP11 constructs were established as described above and maintained in McCoy's 5a Medium Modified, ATCC Catalog No. 30-2007 containing 10% FCS and 400 μg/ml G418 (Geneticin) at 37° C. under an atmosphere of 5% CO2/95% air. Cellular enzyme assays were performed in DMEM/F12 medium containing 2.5% charcoal treated FCS and appropriate concentration of substrate (0.3-10 μM 11-Deoxycorticosterone, 11-Deoxycortisol or Corticosterone). For assaying enzymatic activity, cells were plated onto 96 well plates and incubated for 16 h. An aliquot of the supernatant is then transferred and analyzed for the concentration of the expected product (Aldosterone for CYP11B2; Cortisol for CYP11B1). The concentrations of these steroids can be determined using HTRF assays from CisBio analyzing either Aldosterone or Cortisol.
Measurement: EC50 = 7173.5 nM (pChEMBL 5.139999866485596)
Descriptors: MW 306.37, ALogP 3.18, HBA 3, HBD 0, PSA 50.27, QED 0.81
Compound (SMILES): COc1cc2c(Nc3ccc(Cl)c(Cl)c3)ncnc2cc1OCc1nc2c(s1)CNCC2
InChIKey: WVCAYXOTVLLXMU-UHFFFAOYSA-N
Target: Ephrin type-A receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29317
Assay: Inhibition of recombinant human N-terminal His6-tagged EphA2 (N598 to R890 residues) expressed in baculovirus using poly-EY as substrate after 180 mins by luciferase coupled chemiluminescence assay
Measurement: IC50 = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 488.4, ALogP 5.37, HBA 8, HBD 2, PSA 81.19, QED 0.38
Compound (SMILES): CCCCOC(=O)c1cc(Nc2cc(C3CC3)[nH]n2)nc(NCc2cc(-c3ccccc3)no2)n1
InChIKey: NQVSZVXZRIZPAY-UHFFFAOYSA-N
Target: Insulin-like growth factor 1 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08069
Assay: Inhibition of human IGF1R kinase domain (M954 to C1367 residues) tri-phosphorylated enzyme form expressed in baculovirus based expression system using poly-EY peptide incubated for 2 hrs by luciferase-luciferin coupled chemiluminescent assay
Measurement: IC50 = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 473.54, ALogP 5.04, HBA 9, HBD 3, PSA 130.85, QED 0.2