lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Cc1cc(C)n(C[C@H]2CC[C@H](NC(=O)c3cc(Cl)cnc3C)CC2)n1
InChIKey: FZPRQRQELHONGW-JCNLHEQBSA-N
Target: Corticotropin-releasing factor receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P34998
Assay: In Vitro Inhibition Assay: Chinese hamster ovary (CHO) cells expressing either the human recombinant CRF-1 or CRF-2a receptors (Chen et al., Proc Natl Acad Sci USA 90, 8967-8971, 1993; Liaw et al., Endocrinology 137, 72-77, 1996) are propagated in Dulbecco's modified Eagle medium supplemented with 10% foetal calf serum, non-essential amino acids, 100 U/ml penicillin, 100 mg/l streptomycin and 1 g/l geneticin (G418). For cyclic AMP determinations the Homogeneous Time-Resolved Fluoresce (HTRF) cAMP dynamic 2 kit (Cisbio International, France) was used as per manufacturers' instructions. CHO cells, previously cryopreserved, were thawed, centrifuged for 7 mins at 1200 rpm and resuspended in serum free media, then pipetted out onto clear bottomed black tissue culture treated 384-well microtitre plates (Corning Inc, US) at 2,000 cells per well. Compounds of the invention, prepared in DMSO, and subsequently diluted 50 fold in assay buffer (1x Hanks balanced salt solution, 0.2% (w/v) bovine serum albumin.
Measurement: IC50 = 350 nM (pChEMBL 6.460000038146973)
Descriptors: MW 360.89, ALogP 3.85, HBA 4, HBD 1, PSA 59.81, QED 0.9
Compound (SMILES): Cc1nn(C2CCCCC2)c2sc(C(=O)Nc3ccc(N4CCC(O)CC4)c(F)c3)cc12
InChIKey: QYHFUBZVWDZWRK-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 = 1600 nM (pChEMBL 5.800000190734863)
Descriptors: MW 456.59, ALogP 5.26, HBA 6, HBD 2, PSA 70.39, QED 0.56
Compound (SMILES): C[C@@H](Oc1cc(-c2ccccn2)cc2ncccc12)[C@H]1CNC(=O)C1
InChIKey: JFWWGRGDYNSADT-UKRRQHHQSA-N
Target: Glycogen synthase kinase-3 beta (SINGLE PROTEIN), Homo sapiens
UniProt: P49841
Assay: GSK Kinase Assay: Human GSK3beta (expressed and prified from SF21 cells) is obtained from the University Dundee/Scotland (Dr. James Hastie—Dept. of Biochemistry) in 50 mM Tris (pH7.5); 150 mM NaCl; 0.1 mM EGTA, 270 mM Succrose, 0.1% β-mercaptoethanol, 1 mM benzamidine, 0.2 mM PMSF; sequence see below). The enzyme is diluted to 3.56 μM (168 μg/ml) in enzyme dilution buffer and 6 μl aliquots are stored at −80° C.The activity of GSK3 kinase protein is measured using the Z′-LYTETM assay technology from Invitrogen (# PV3324).
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 333.39, ALogP 3.2, HBA 4, HBD 1, PSA 64.11, QED 0.8
Compound (SMILES): CN(C)[C@@H]1CCC(n2cc(C(N)=O)c(Nc3ccc(S(=O)(=O)C(F)(F)F)cc3)n2)[C@@H](C#N)C1
InChIKey: WQPOTWFXOOAHHW-VDCCYECJSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: HTRF Assay: The ability of compounds to inhibit the activity of JAK1, JAK2, JAK3, and Tyk2 was measured using a recombinant purified GST-tagged catalytic domain for each enzyme (Invitrogen JAK1 #M4290, JAK2 #M4290, JAK3 #M4290, Tyk2 #M4290) in an HTRF format biochemical assay. The reactions employed a common peptide substrate, LCB-EQEDEPEGDYFEWLW-NH2 (in-house). The basic assay protocol is as follows: First, 250 mL of diluted compounds in DMSO were dispensed into the wells of a dry 384-well Black plate (Greiner #781076) using a Labcyte Echo 555 acoustic dispenser. Subsequent reagent additions employed an Agilent Bravo. Next, 18 uL of 1.11x enzyme and 1.11x substrate in 1x assay buffer (Invitrogen kinase buffer #PV3189, 2 mM DTT, 0.05% BSA) were added to the wells and shaken and then preincubated for 30 minutes at ambient temperature to allow compound binding to equilibrate. After equilibration, 2 uL of 10xATP in 1x assay buffer was added to initiate the kinase reaction.
Measurement: IC50 = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 484.5, ALogP 2.81, HBA 8, HBD 2, PSA 134.11, QED 0.64
Compound (SMILES): N#C[C@H]1CCC2(C[C@@H]1n1cc(C(N)=O)c(Nc3ccccc3)n1)OCCO2
InChIKey: GOYVGGAPXFQVDR-CJNGLKHVSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: HTRF Assay: The ability of compounds to inhibit the activity of JAK1, JAK2, JAK3, and Tyk2 was measured using a recombinant purified GST-tagged catalytic domain for each enzyme (Invitrogen JAK1 #M4290, JAK2 #M4290, JAK3 #M4290, Tyk2 #M4290) in an HTRF format biochemical assay. The reactions employed a common peptide substrate, LCB-EQEDEPEGDYFEWLW-NH2 (in-house). The basic assay protocol is as follows: First, 250 mL of diluted compounds in DMSO were dispensed into the wells of a dry 384-well Black plate (Greiner #781076) using a Labcyte Echo 555 acoustic dispenser. Subsequent reagent additions employed an Agilent Bravo. Next, 18 uL of 1.11x enzyme and 1.11x substrate in 1x assay buffer (Invitrogen kinase buffer #PV3189, 2 mM DTT, 0.05% BSA) were added to the wells and shaken and then preincubated for 30 minutes at ambient temperature to allow compound binding to equilibrate. After equilibration, 2 uL of 10xATP in 1x assay buffer was added to initiate the kinase reaction.
Measurement: IC50 = 0.85 nM (pChEMBL 9.069999694824219)
Descriptors: MW 367.41, ALogP 2.33, HBA 7, HBD 2, PSA 115.19, QED 0.86
Compound (SMILES): NC1=NC2(CO1)c1cc(NC(=O)c3ncc(Br)cn3)ccc1OCC21CC1
InChIKey: RUPHOSKJXJYHAD-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 43 nM (pChEMBL 7.369999885559082)
Descriptors: MW 430.26, ALogP 2.2, HBA 7, HBD 2, PSA 111.72, QED 0.76
Compound (SMILES): CC1(C)Oc2ccc(NC(=O)c3ncc(Br)cn3)cc2C2(COC(N)=N2)C12COC2
InChIKey: OKNLKKUCXCACGI-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 4.1 nM (pChEMBL 8.390000343322754)
Descriptors: MW 474.32, ALogP 2.22, HBA 8, HBD 2, PSA 120.95, QED 0.68
Compound (SMILES): Cc1cc(C#N)cnc1C(=O)Nc1ccc2c(c1)C1(COC(N)=N1)C1(CC1)CO2
InChIKey: LUKKVICWYPMUPG-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 27 nM (pChEMBL 7.570000171661377)
Descriptors: MW 389.42, ALogP 2.23, HBA 7, HBD 2, PSA 122.62, QED 0.81
Compound (SMILES): N#Cc1cnc(C(=O)Nc2ccc3c(c2)C2(COC(N)=N2)C2(CC2)CO3)c(Cl)c1
InChIKey: OBYNGUKQVYJXLG-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 30 nM (pChEMBL 7.519999980926514)
Descriptors: MW 409.83, ALogP 2.57, HBA 7, HBD 2, PSA 122.62, QED 0.79
Compound (SMILES): Cc1cc(Cl)cnc1C(=O)Nc1ccc2c(c1)C1(COC(N)=N1)C1(CC1)CO2
InChIKey: UGPKGDXAPPRVIR-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 28 nM (pChEMBL 7.550000190734863)
Descriptors: MW 398.85, ALogP 3.01, HBA 6, HBD 2, PSA 98.83, QED 0.81
Compound (SMILES): Cc1cc(Cl)cnc1C(=O)Nc1ccc2c(c1)C1(COC(N)=N1)C1(COC1)C(C)(C)O2
InChIKey: WTTPTTMLIGTSRC-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 3.2 nM (pChEMBL 8.489999771118164)
Descriptors: MW 442.9, ALogP 3.02, HBA 7, HBD 2, PSA 108.06, QED 0.74
Compound (SMILES): N#Cc1ccc(C(=O)Nc2ccc3c(c2)C2(COC(N)=N2)C2(CC2)CO3)nc1
InChIKey: FCSDLAQZROWWFP-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 53 nM (pChEMBL 7.28000020980835)
Descriptors: MW 375.39, ALogP 1.92, HBA 7, HBD 2, PSA 122.62, QED 0.83
Compound (SMILES): CC1(C)Oc2ccc(NC(=O)c3ccc(C#N)cn3)cc2C2(COC(N)=N2)C12COC2
InChIKey: GPWZMCGYHYETLO-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 5.7 nM (pChEMBL 8.239999771118164)
Descriptors: MW 419.44, ALogP 1.93, HBA 8, HBD 2, PSA 131.85, QED 0.76
Compound (SMILES): NC1=NC2(CO1)c1cc(-c3cncnc3)ccc1OC(Cc1ccccc1)C21COC1
InChIKey: STMXAUVFLJMWDN-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Fluorescence Resonance Energy Transfer (FRET) Assay: Potency of test compounds were determined by measurement of their inhibition of BACE1 activity toward a fluorescent substrate. Experiments were performed by reference to the procedure as described in Ermolieff, et al. (Biochemistry 39:12450-12456 (2000), the teachings of which are incorporated hereby in their entirety). Briefly, the recombinant protease unit of BACE1 was prepared from E. coli expression as inclusion bodies, refolded, and purified as described in Lin, et al., (Proc. Nat. Acad. Sci. 97:1456-1460 (2000)). Fluorogenic substrate, MCA-SEVNLDAEFK(DNP)-NH2 (SEQ ID NO:1) was purchased. (M-2485, Bachem Americas, Torrance, Calif.). The substrate was derived from 10 amino acids of the human amyloid precursor protein (APP), with the Swedish variant amino acids at the beta-secretase cleavage site. The terminal amino acid was modified from arginine to lysine to facilitate derivatization with a functional group for detection by autofluorescence.
Measurement: IC50 = 145 nM (pChEMBL 6.840000152587891)
Descriptors: MW 414.47, ALogP 2.7, HBA 7, HBD 1, PSA 91.85, QED 0.71
Compound (SMILES): CC#Cc1cncc(-c2ccc3c(c2)C2(COC(N)=N2)C2(COC2)C(Cc2ccccc2)O3)c1
InChIKey: UZRCBYZWOAYXCE-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Fluorescence Resonance Energy Transfer (FRET) Assay: Potency of test compounds were determined by measurement of their inhibition of BACE1 activity toward a fluorescent substrate. Experiments were performed by reference to the procedure as described in Ermolieff, et al. (Biochemistry 39:12450-12456 (2000), the teachings of which are incorporated hereby in their entirety). Briefly, the recombinant protease unit of BACE1 was prepared from E. coli expression as inclusion bodies, refolded, and purified as described in Lin, et al., (Proc. Nat. Acad. Sci. 97:1456-1460 (2000)). Fluorogenic substrate, MCA-SEVNLDAEFK(DNP)-NH2 (SEQ ID NO:1) was purchased. (M-2485, Bachem Americas, Torrance, Calif.). The substrate was derived from 10 amino acids of the human amyloid precursor protein (APP), with the Swedish variant amino acids at the beta-secretase cleavage site. The terminal amino acid was modified from arginine to lysine to facilitate derivatization with a functional group for detection by autofluorescence.
Measurement: IC50 = 109 nM (pChEMBL 6.960000038146973)
Descriptors: MW 451.53, ALogP 3.68, HBA 6, HBD 1, PSA 78.96, QED 0.61
Compound (SMILES): CC#Cc1cncc(-c2ccc3c(c2)C2(COC(N)=N2)C2(COC2)C(CC2CC2)O3)c1
InChIKey: KJZOKUPYIWIFKS-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 22 nM (pChEMBL 7.659999847412109)
Descriptors: MW 415.49, ALogP 3.24, HBA 6, HBD 1, PSA 78.96, QED 0.78
Compound (SMILES): CC1(C)Oc2ccc(NC(=O)c3ccc(Cl)cn3)cc2[C@@]2(COC(N)=N2)C12COC2
InChIKey: INWIVMOTHPFWNU-NRFANRHFSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Fluorescence Resonance Energy Transfer (FRET) Assay: Potency of test compounds were determined by measurement of their inhibition of BACE1 activity toward a fluorescent substrate. Experiments were performed by reference to the procedure as described in Ermolieff, et al. (Biochemistry 39:12450-12456 (2000), the teachings of which are incorporated hereby in their entirety). Briefly, the recombinant protease unit of BACE1 was prepared from E. coli expression as inclusion bodies, refolded, and purified as described in Lin, et al., (Proc. Nat. Acad. Sci. 97:1456-1460 (2000)). Fluorogenic substrate, MCA-SEVNLDAEFK(DNP)-NH2 (SEQ ID NO:1) was purchased. (M-2485, Bachem Americas, Torrance, Calif.). The substrate was derived from 10 amino acids of the human amyloid precursor protein (APP), with the Swedish variant amino acids at the beta-secretase cleavage site. The terminal amino acid was modified from arginine to lysine to facilitate derivatization with a functional group for detection by autofluorescence.
Measurement: IC50 = 30300 nM (pChEMBL 4.519999980926514)
Descriptors: MW 428.88, ALogP 2.72, HBA 7, HBD 2, PSA 108.06, QED 0.76
Compound (SMILES): CC1(C)Oc2ccc(NC(=O)c3ccc(Br)cn3)cc2[C@@]2(COC(N)=N2)C12COC2
InChIKey: CWCLNAMDVBTHAU-NRFANRHFSA-N
Target: Voltage-gated inwardly rectifying potassium channel KCNH2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q12809
Assay: Inhibition Assay: The hERG potassium current was measured in a hERG-stably-expressing Chinese hamster ovary K1 (CHO) cells. The experiments were performed using an automated planar patch-clamp system QPatch HTX (Sophion Bioscience A/S). The application of pressure for forming gigaseals and whole-cell patch clamp configuration were established using the QPatch assay software. Patch-clamp experiments were performed in voltage-clamp mode and whole-cell currents were recorded. The following stimulation protocol was applied to investigate the effects of compounds on hERG potassium channel.The membrane potential was held at −80 mV and repetitively (every 15 seconds) depolarized to +20 mV for 4800 milliseconds after the pulse to −50 mV for 20 milliseconds served to define the baseline, followed by repolarizing step to −50 mV for 5000 milliseconds to evaluate of the tail current amplitude. Experiments were conducted at room temperature.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 473.33, ALogP 2.82, HBA 7, HBD 2, PSA 108.06, QED 0.69
Compound (SMILES): COc1cnc(C(=O)Nc2ccc3c(c2)[C@@]2(COC(N)=N2)C2(COC2)C(C)(C)O3)cn1
InChIKey: WZWNQLAVZZUSHH-NRFANRHFSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 3.1 nM (pChEMBL 8.510000228881836)
Descriptors: MW 425.45, ALogP 1.47, HBA 9, HBD 2, PSA 130.18, QED 0.76
Compound (SMILES): CC1(C)Oc2ccc(NC(=O)c3ccc(Cl)cn3)cc2[C@@]2(COC(N)=N2)C12CC2
InChIKey: SUARPGUFOBBSJS-NRFANRHFSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 0.16 nM (pChEMBL 9.800000190734863)
Descriptors: MW 412.88, ALogP 3.48, HBA 6, HBD 2, PSA 98.83, QED 0.79
Compound (SMILES): CC1(C)Oc2ccc(NC(=O)c3ccc(F)cn3)cc2[C@@]2(COC(N)=N2)C12CC2
InChIKey: KFVQLLMGMYDUOM-NRFANRHFSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 0.62 nM (pChEMBL 9.210000038146973)
Descriptors: MW 396.42, ALogP 2.96, HBA 6, HBD 2, PSA 98.83, QED 0.81
Compound (SMILES): COc1cnc(C(=O)Nc2ccc3c(c2)[C@@]2(COC(N)=N2)C2(CC2)C(C)(C)O3)cn1
InChIKey: FVLGCKAZTKQUNT-NRFANRHFSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 3.1 nM (pChEMBL 8.510000228881836)
Descriptors: MW 409.45, ALogP 2.23, HBA 8, HBD 2, PSA 120.95, QED 0.8
Compound (SMILES): CC1(C)Oc2ccc(NC(=O)c3cnc(C(F)F)cn3)cc2[C@@]2(COC(N)=N2)C12CC2
InChIKey: UUOAKQJQRFDOFS-NRFANRHFSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 0.63 nM (pChEMBL 9.199999809265137)
Descriptors: MW 429.43, ALogP 3.16, HBA 7, HBD 2, PSA 111.72, QED 0.78
Compound (SMILES): COc1cnc(C(=O)Nc2ccc3c(c2)C2(COC(N)=N2)C2(COC2)C(C)(C)O3)cn1
InChIKey: WZWNQLAVZZUSHH-UHFFFAOYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: Cellular Assay of Aβ Production: Test Example 3: The potency of compounds against BACE1 activity was determined in a cellular assay of Aβ production. Human SK-N-BE(2) neuroblastoma cells (ATCC No. CRL-2271) were plated at 96,000 cells/well/100 uL in 96-well plates in RPMI1640 medium/10% fetal bovine serum (FBS)/penicillin-streptomycin and cultured for 24 hours at 37° C., 5% CO2. Test compounds were dissolved in dimethyl sulfoxide and diluted with dimethyl sulfoxide and put into RPMI1640/5% FBS/penicillin-streptomycin media (final dimethyl sulfoxide concentration is 0.5%). The culture media in 96-well plates were replaced by 125 uL/well of the media containing test compounds. After incubation for 6 hours at 37° C., 5% CO2, 30 uL of the media were transferred into a fresh 96-well plate and used for Aβ40 assay by an enzyme-linked immunosorbent assay (ELISA) kit (#27718, Immuno-Biological Laboratories, Japan) according to the manufacturer's protocol. Cell viability was measured by CellTiter-Glo Luminescent Cell Viability Assay (#7571, Promega) after removal of 30 uL of the media for the Aβ assay.
Measurement: IC50 = 8 nM (pChEMBL 8.100000381469727)
Descriptors: MW 425.45, ALogP 1.47, HBA 9, HBD 2, PSA 130.18, QED 0.76
Compound (SMILES): Cc1[nH]nc2c1C(c1ccc(Cl)cc1)N(c1cc(Cl)c(=O)n(C)c1)C2=O
InChIKey: ZMFIMBNRJYXBHU-UHFFFAOYSA-N
Target: Bromodomain-containing protein 3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q15059
Assay: TR-FRET In-Vitro Binding Assay: All assays were performed in 384-well microtiter plates. Each assay plate contained 8-point serial dilutions for 40 test compounds, plus 16 high- and 16 low controls. Liquid handling and incubation steps were done on an Innovadyne Nanodrop Express equipped with a robotic arm (Thermo CatX, Perkin Elmer/Caliper Twister II) and an incubator (Liconic STX40, Thermo Cytomat 2C450). The assay plates were prepared by addition of 50 nl per well of compound solution in 90% DMSO HummingBird nanodispenser (Zinsser Analytic). The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.1% BSA, 50 mM NaCl, 45 nM His-Brd2(60-472) or 45 nM His-Brd3(20-477) or 45 nM His-Brd4(44-477) all proteins produced in-house) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.1% BSA, 50 mM NaCl, 60 nM acetyl-histone H4 (AcK 5, 8, 12, 16) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes.
Measurement: IC50 = 370 nM (pChEMBL 6.429999828338623)
Descriptors: MW 389.24, ALogP 3.47, HBA 4, HBD 1, PSA 70.99, QED 0.73
Compound (SMILES): Cc1c2c(nn1C)C(=O)N(c1cc(Cl)c(=O)n(C)c1)C2c1ccc(Cl)cc1
InChIKey: SSCBXXKTZLTDDN-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 3500 nM (pChEMBL 5.460000038146973)
Descriptors: MW 403.27, ALogP 3.48, HBA 5, HBD 0, PSA 60.13, QED 0.66
Compound (SMILES): Cc1c2c(nn1C1CC1)C(=O)N(c1cc(Cl)c(=O)n(C)c1)C2c1ccc(Cl)cc1
InChIKey: FTYXFBKWRBABJZ-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 1300 nM (pChEMBL 5.889999866485596)
Descriptors: MW 429.31, ALogP 4.28, HBA 5, HBD 0, PSA 60.13, QED 0.62
Compound (SMILES): Cc1cc(N2C(=O)c3nn(C4CC4)c(C)c3C2c2ccc(Cl)cc2)cn(C)c1=O
InChIKey: FTHLNVOKHACXIU-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 1100 nM (pChEMBL 5.960000038146973)
Descriptors: MW 408.89, ALogP 3.94, HBA 5, HBD 0, PSA 60.13, QED 0.66
Compound (SMILES): Cc1nn(C2CC2)c2c1C(c1ccc(Cl)cc1)N(c1cc(Cl)c(=O)n(C)c1)C2=O
InChIKey: KILRLNJOBPEYAV-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 5900 nM (pChEMBL 5.230000019073486)
Descriptors: MW 429.31, ALogP 4.28, HBA 5, HBD 0, PSA 60.13, QED 0.62
Compound (SMILES): Cc1[nH]nc2c1C(c1ccc(Cl)cc1)N(c1cc(C)c(=O)n(C)c1)C2=O
InChIKey: GXKIVKDALCGWBS-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 3800 nM (pChEMBL 5.420000076293945)
Descriptors: MW 368.82, ALogP 3.13, HBA 4, HBD 1, PSA 70.99, QED 0.76
Compound (SMILES): Cc1cc(N2C(=O)c3nn(C)c(C)c3C2c2ccc(Cl)cc2)cn(C)c1=O
InChIKey: QKILPYXMYOTNIL-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 3300 nM (pChEMBL 5.480000019073486)
Descriptors: MW 382.85, ALogP 3.14, HBA 5, HBD 0, PSA 60.13, QED 0.68
Compound (SMILES): Cc1noc2ccc(N3C(=O)c4nn(C)c(C)c4C3c3ccc(Cl)cc3)cc12
InChIKey: KKNOUVNOCNNTDD-UHFFFAOYSA-N
Target: Bromodomain-containing protein 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P25440
Assay: TR-FRET In-Vitro Binding Assay: All assays were performed in 384-well microtiter plates. Each assay plate contained 8-point serial dilutions for 40 test compounds, plus 16 high- and 16 low controls. Liquid handling and incubation steps were done on an Innovadyne Nanodrop Express equipped with a robotic arm (Thermo CatX, Perkin Elmer/Caliper Twister II) and an incubator (Liconic STX40, Thermo Cytomat 2C450). The assay plates were prepared by addition of 50 nl per well of compound solution in 90% DMSO HummingBird nanodispenser (Zinsser Analytic). The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.1% BSA, 50 mM NaCl, 45 nM His-Brd2(60-472) or 45 nM His-Brd3(20-477) or 45 nM His-Brd4(44-477) all proteins produced in-house) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.1% BSA, 50 mM NaCl, 60 nM acetyl-histone H4 (AcK 5, 8, 12, 16) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes.
Measurement: IC50 = 31200 nM (pChEMBL 4.510000228881836)
Descriptors: MW 392.85, ALogP 4.58, HBA 5, HBD 0, PSA 64.16, QED 0.5
Compound (SMILES): Cc1cc(N2C(=O)c3c(cnn3C3CC3)C2c2ccc(Cl)cc2)cn(C)c1=O
InChIKey: MWVVALQFXSOQLF-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 5200 nM (pChEMBL 5.28000020980835)
Descriptors: MW 394.86, ALogP 3.63, HBA 5, HBD 0, PSA 60.13, QED 0.68
Compound (SMILES): Cn1cc(N2C(=O)c3c(cnn3C3CC3)C2c2ccc(Cl)cc2)cc(Cl)c1=O
InChIKey: DDCDPEKGXFRTDS-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 7400 nM (pChEMBL 5.130000114440918)
Descriptors: MW 415.28, ALogP 3.97, HBA 5, HBD 0, PSA 60.13, QED 0.65
Compound (SMILES): Cc1cc(N2C(=O)c3c(cnn3C(C)C)C2c2ccc(Cl)cc2)cn(C)c1=O
InChIKey: ZNGYYGUCWRDEAS-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 3600 nM (pChEMBL 5.440000057220459)
Descriptors: MW 396.88, ALogP 3.87, HBA 5, HBD 0, PSA 60.13, QED 0.67
Compound (SMILES): Cn1cc(N2C(=O)c3nn(C4CC4)cc3C2c2ccc(Cl)cc2)cc(Cl)c1=O
InChIKey: RQSRLTLIMDDPDM-UHFFFAOYSA-N
Target: CREB-binding protein (SINGLE PROTEIN), Homo sapiens
UniProt: Q92793
Assay: AlphaScreen In-Vitro Binding Assay: In order to assess bromodomain selectivity, we set up a binding assay using the bromodomain encoded by the CREBBP gene. Compounds were tested in the CREBBP assay with a similar protocol, however using AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay, Perkin Elmer) as detection readout instead of TR-FRET. The assay was started by stepwise addition of 4.5 uL per well of bromo domain protein (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 324 nM His-CREBBP (1081-1197) (custom production at Viva Biotech Ltd.)) and 4.5 uL per well of peptide solution (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 120 nM acetyl-histone H4 (AcK 5, 8, 12) (Biosyntan GmbH)). Reactions were incubated at 30 C. for 35 minutes. Subsequently 4.5 uL per well detection mix (50 mM HEPES, pH 7.5, 0.005% Tween20, 0.02% BSA, 150 mM NaCl, 45 ug/ml Ni-chelate acceptor beads, 45 ug/mL streptavidin-donor beads) (Perkin Elmer)) were added.
Measurement: IC50 = 1200 nM (pChEMBL 5.920000076293945)
Descriptors: MW 415.28, ALogP 3.97, HBA 5, HBD 0, PSA 60.13, QED 0.65