lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): CC(C(N)=O)c1ccccc1CCc1nc(Nc2cnn(C3CCN(C)CC3)c2)ncc1C(F)(F)F
InChIKey: RJUAJPXDDBGVQP-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P35916
Assay: VEGFR3 Phospho ELISA Assay: Adult human dermal lymphatic microvascular endothelial cells (HMVEC-dLyAD) (Cat# CC-2810, Lonza) were seeded into clear-bottom, TC treated 12 well plates (Cat #665180, Greiner Bio-One) in EGM-2MV (Cat# CC-3202, Lonza) at 180,000 cells/well (volume 1 mL), and the plates incubated at 37° C. and 5% CO2 for 6 hours. The media was replaced with EBM-2 (Cat # CC-3156, Lonza)+0.1% BSA (Cat# A8412, Sigma) and cells incubated for a further period (overnight at 37° C. and 5% CO2).96 well Maxisorp immuno plates (Cat #439454, Nunc) were coated with 100 uL of Total VEGFR3 capture antibody (Part #841888, Human Total VEGFR3/FLT4 ELISA Kit, Cat # DYC3491, R&D Systems), or Phospho VEGFR3 Capture antibody (Part #841885, Human Phospho VEGFR3/FLT4 ELISA Kit, Cat# DYC2724, R&D Systems). The plates were covered and incubated at room temperature overnight .The coating antibody was flicked out and the plates washed three times with Wash Buffer (Phosphate buffered saline (137 mM NaCl, 2.7 nM KCl, 8.1 nM Na2HPO4, 1.5 mL KH2PO4, pH 7.2-7.4), 0.05% Tween 20). 300 uL of blocking buffer (5% v/v Tween 20, 5% w/v sucrose in PBS) was then added to wells and plate incubated for 2 hours at room temperature.
Measurement: IC50 = 27 nM (pChEMBL 7.570000171661377)
Descriptors: MW 501.56, ALogP 4.08, HBA 7, HBD 2, PSA 101.96, QED 0.48
Compound (SMILES): N=C(N)NCCCCC(=O)Nc1ccc(C[C@H](NC(=O)[C@@H]2CCN2S(=O)(=O)c2ccccc2)C(=O)O)cc1
InChIKey: WJYNGTHNURVQMC-SFTDATJTSA-N
Target: Integrin alpha-V/beta-1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P06756
Assay: Inhibition of integrin alphaVbeta1 (unknown origin)-mediated CHO cell adhesion to fibronectin preincubated for 15 to 30 mins followed by 60 min incubation for adhesion by crystal violet staining based cell adhesion assay
Measurement: IC50 = 160 nM (pChEMBL 6.800000190734863)
Descriptors: MW 544.63, ALogP 0.85, HBA 6, HBD 6, PSA 194.78, QED 0.12
Compound (SMILES): O=C(NCc1cccc(-c2ccc(CS(=O)(=O)Cc3n[nH]c(=O)[nH]3)cc2)c1)c1nc2scc(-c3cccc(Cl)c3)c2c(=O)[nH]1
InChIKey: HGMNWCDIIMBGAR-UHFFFAOYSA-N
Target: Stromelysin-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P09238
Assay: Inhibition of recombinant human AMPA-activated MMP10 using Cy3-PLGLK(Cy5Q)AR-NH2 as substrate measured after 40 mins by spectrofluorimetric method
Measurement: IC50 = 34 nM (pChEMBL 7.46999979019165)
Descriptors: MW 647.14, ALogP 4.43, HBA 8, HBD 4, PSA 170.53, QED 0.18
Compound (SMILES): CC(C)CC(=O)N(Cc1ccccc1)[C@H](C)CN1[C@@H]2CC[C@H]1C[C@@H](c1cccc(O)c1)C2
InChIKey: JQNMPIPXXJPAFY-ATPOPXNDSA-N
Target: Delta-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41143
Assay: Displacement of [3H]-diprenorphine from human delta opioid receptor expressed in cell membranes
Measurement: Ki = 100 nM (pChEMBL 7)
Descriptors: MW 434.62, ALogP 5.57, HBA 3, HBD 1, PSA 43.78, QED 0.59
Compound (SMILES): c1ccc2c(c1)ncn2-c1cc(Nc2ccc(OCCN3CCOCC3)cn2)ncn1
InChIKey: REJHOMMDDHCCJU-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 2/cyclin E1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P24864
Assay: Inhibition of recombinant full-length human C-terminal His6-tagged CDK2/recombinant full-length human N-terminal GST-tagged cyclin E using fluorescein-QSPKKG-CONH2 as substrate after 60 mins by off-chip mobility shift assay
Measurement: IC50 = 554 nM (pChEMBL 6.260000228881836)
Descriptors: MW 417.47, ALogP 2.67, HBA 9, HBD 1, PSA 90.22, QED 0.49
Compound (SMILES): Cc1cc([C@](C)(O)C#Cc2ccc3c(c2)-c2nc(C(N)=O)cn2C[C@@H](C(C)(C)O)O3)no1
InChIKey: RMSCKOLYCLRFFV-WMZHIEFXSA-N
Target: Mitogen-activated protein kinase kinase kinase 14 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99558
Assay: Inhibition Assay: The ability of the nuclear factor-kappa B (NF-kB)-inducing kinase (NIK) to catalyze the hydrolysis of adenosine-5'-triphosphate (ATP) was monitored using the Transcreener ADP (adenosine-5'-diphosphate) assay (BellBrook Labs). Purified NIK (0.2-1 nM) derived from a baculovirus-infected insect cell expression system was incubated with test compounds for 1-3.5 hours in 50 mM 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid buffer (pH 7.2) containing 10 mM MgCl.sub.2, 2 mM dithiothreitol, 10 uM ATP, 0.01% Triton X-100, 0.1% gamma-globulins from bovine blood, 1% dimethylsulfoxide (DMSO), 12 ug/mL ADP antibody and 4 nM ADP-AlexaFluor.RTM. 633 tracer. Reactions were quenched by the addition of 20 mM 2,2',2'',2'''-(ethane-1,2-diyldinitrilo)tetraacetic acid and 0.01% Brij 35. The tracer bound to the antibody was displaced by the ADP generated during the NIK reaction, which causes a decrease in fluorescence polarization that was measured by laser excitation.
Measurement: Ki = 250 nM (pChEMBL 6.599999904632568)
Descriptors: MW 436.47, ALogP 1.74, HBA 8, HBD 3, PSA 136.63, QED 0.53
Compound (SMILES): Cc1cc(Oc2ccnc(NC(=O)C3CC3)c2)ccc1NC(=O)Nc1ccc(F)c(C(F)(F)F)c1
InChIKey: GVMPOIUFFPTGSJ-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition Assay: [Experimental method]: Use ADP-GLO™ assay kit from Promega to evaluate the inhibitory activity of compounds at B-Raf. [Instrument]: PerKinElmer's ENVISION plate reader. [Material]: Optiplate-384 well plate (PerKinElmer), kinase buffer (50 mM Hepes pH7.5, 1 mM EGTA, 2 mM DTT, 10 mM Mg2+, 0.05% BSA), B-Raf kinase (Millipore, catalogue #14-530-K), GST-MEK1 substrate (Carna, catalogue #07-141-10), Super pure ATP (Promega), ADP-GLO™ assay kit (Promega, catalogue # V9102), DMSO (Sigma, catalogue #34869), purified water (Millipore, type: Milli-Q). [Study conditions]: Mix B-Raf kinase (final concentration: 5 nM) and compound (DMSO final concentration: 0.25%) and preincubate at 30° C. for 20 min, then add ATP (final concentration: 6 μM) and reaction substrate (final concentration: 30 nM), React for 2 hrs at 30° C. After reaction, add ADP-GLO™ reagent, react for 40 min at room temperature, then add ADP-GLO™ detecting agent, react for 30 min at room temperature. After that, use ENVSION to measure the fluorescence.
Measurement: IC50 = 10 nM (pChEMBL 8)
Descriptors: MW 488.44, ALogP 6.33, HBA 4, HBD 3, PSA 92.35, QED 0.35
Compound (SMILES): N[C@@H]1[C@H]2CN(c3ncc(C(=O)Nc4ccc(OC(F)(F)Cl)cc4)cc3-c3cncnc3)C[C@@H]12
InChIKey: SLPHANANQOBVQJ-BWTSREIZSA-N
Target: Tyrosine-protein kinase ABL1 (SINGLE PROTEIN), Homo sapiens
UniProt: P00519
Assay: Caliper ABL : 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 uL 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 uM sodium orthovanadate, 20 mM MgCl2, 2 mM MnCl2, 4 uM ATP, 4 uM peptide (FITC-Ahx-EAIYAAPFAKKK-NH2)) and 4.5 uL 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 uM 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 uL 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.
Measurement: IC50 = 0.5 nM (pChEMBL 9.300000190734863)
Descriptors: MW 472.88, ALogP 3.35, HBA 7, HBD 2, PSA 106.26, QED 0.53
Compound (SMILES): Cc1ccc(C2=C(C(=O)NC3CC3)C(=O)NC(c3ccc(OCCCC(F)(F)F)cc3)(C(F)(F)F)C2)cc1
InChIKey: AGVPRSKJBAHSEV-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 = 94 nM (pChEMBL 7.03000020980835)
Descriptors: MW 540.5, ALogP 5.73, HBA 3, HBD 2, PSA 67.43, QED 0.26
Compound (SMILES): CN[C@@H](C)C(=O)N[C@H]1C(=O)N(Cc2c(OC)ccc3cc(Br)ccc23)c2ccccc2O[C@@]12CCOC2
InChIKey: HELQGTDTITZVDX-JGXREDOWSA-N
Target: E3 ubiquitin-protein ligase XIAP (SINGLE PROTEIN), Homo sapiens
UniProt: P98170
Assay: TR-FRET Assay: Ten nanomolar of 6x Histidine-tagged BIR2 domain, corresponding to amino acids 124-240 of XIAP, or BIR3 domain, corresponding to amino acids 241-356 of XIAP, was mixed with 20 nM of the peptide AVPIAQKSEK-(8-biotin)-OH 1:2 TFA, in the presence of 50 mM Tris-C1, pH 7.5, 100 mM NaCl, 1 mM dithiothreitol (DTT) and 0.1 mg/mL bovine serum albumin (BSA). Following a 45 min. incubation at 37° C., Europium-Streptavidin and Allophycocyanin conjugated anti-Histidine antibody were added to a final concentration of 1.5 nM and 15 nM, respectively. Time-resolved fluorescence resonance energy transfer (TR-FRET) signals were measured 1 hour later at room temperature. Test compound potency was assessed at 10 serially diluted concentrations.
Measurement: IC50 = 20630 nM (pChEMBL 4.690000057220459)
Descriptors: MW 568.47, ALogP 3.79, HBA 6, HBD 2, PSA 89.13, QED 0.47
Compound (SMILES): CC(CO)n1cc(C(=O)c2cncc(NC(=O)Cc3ccc(F)cn3)c2)c2cnccc21
InChIKey: SYFRPNJQPGCXIP-UHFFFAOYSA-N
Target: High affinity nerve growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P04629
Assay: HTRF Assay: Isolated TRK Enzyme assays use the HTRF KinEASE-TK kit (Cisbio Cat#62TK0PEJ) with recombinant His-tagged cytoplasmic domains of each TRK receptor sourced from Invitrogen (see table below). This activity-assay measures the phosphorylation of tyrosine residues within a substrate from the HTRF kit which has been validated by Cisbio for a variety of tyrosine kinases including the TRK receptors.Assay Details: Invitrogen Amino FAC FAC Assay ReactionTarget Cat# acids enzyme ATP TimeTRKA PV3144 aa 441-4 nM 40 uM 35 min (NTRK1) 796 TRKB PV3616 aa 526-1 nM 1.4 uM 40 min (NTRK2) 838 TRKC PV3617 aa 510- 10 nM 15 uM 30 min (NTRK3) 8250.5 mM stock solutions of test compounds are prepared and serially diluted in 100% DMSO. A standard curve using the compound of Example 135 disclosed in WO2005/116035 of 150 uM is also prepared on each test plate. High percentage effect (HPE) is defined by 150 uM.
Measurement: IC50 = 233 nM (pChEMBL 6.630000114440918)
Descriptors: MW 433.44, ALogP 2.93, HBA 7, HBD 2, PSA 110, QED 0.43
Compound (SMILES): Cc1ccc(-c2cnc(N)c(C(=O)Nc3ccncc3)c2)s1
InChIKey: UEESRKKBPBMNHE-UHFFFAOYSA-N
Target: Inhibitor of nuclear factor kappa-B kinase subunit epsilon (SINGLE PROTEIN), Homo sapiens
UniProt: Q14164
Assay: IKKepsilon Flashplate Assay: The kinase assay is performed either as 384-well Flashplate assay (for e.g. Topcount measurement). 1 nM IKKepsilon, 800 nM biotinylated IkappaBalpha(19-42) peptide (Biotin-C6-C6-GLKKERLLDDRHDSGLDSMKDEE) SEQ ID NO: 1 and 10 uM ATP (spiked with 0.3 uCi 33P-ATP/well) are incubated in a total volume of 50 ul (10 mM MOPS, 10 mM Mg-acetat, 0.1 mM EGTA, 1 mM Dithiothreitol,0.02% Brij35, 0.1% BSA, 0.1% BioStab, pH 7.5) with or without test compound for 2 hours at 30° C. The reaction is stopped with 25 ul 200 mM EDTA. After 30 Min at room temperature the liquid is removed and each well washed thrice with 100 ul 0.9% sodium chloride solution. Non-specific reaction is determined in presence of 3 uM MSC2119074 (BX-795). Radioactivity is measured with Topcount (PerkinElmer).
Measurement: IC50 = 150 nM (pChEMBL 6.820000171661377)
Descriptors: MW 310.38, ALogP 3.35, HBA 5, HBD 2, PSA 80.9, QED 0.78
Compound (SMILES): CN1CCC2(CC1)C(=N)N[C@](C)(c1cc(NC(=O)c3ccc(C#N)cn3)ccc1F)CS2(=O)=O
InChIKey: NYROTPFSKHMAEQ-QFIPXVFZSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: Time-Resolved Endpoint Proteolysis Assay: Inhibitor IC50s at purified human autoBACE-2 are determined in a time-resolved endpoint proteolysis assay that measures hydrolysis of the QSY7-EISEVNLDAEFC-Eu-amide FRET peptide substrate (BACE-HTRF assay). BACE-mediated hydrolysis of this peptide results in an increase in relative fluorescence (RFU) at 620 nm after excitation with 320 nm light. Inhibitor compounds, prepared at 3x the desired final concentration in 1Ã¿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 1xBACE 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 uM for 4 uM for autoBACE-2) prepared in 1xBACE 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 us delay on a RUBYstar HTRF plate reader (BMG Labtechnologies). Raw RFU data is normalized to maximum (1.0 nM BACE/DMSO) and minimum (no enzyme/DMSO) RFU values. IC50s are determined by nonlinear regression analysis (sigmoidal dose response, variable slope) of percent inhibition data with minimum and maximum values set to 0 and 100 percent respectively. Similar IC50s are obtained when using raw RFU data. The Ki values are calculated from the IC50 using the Cheng-Prusoff equation.
Measurement: Ki = 8.4 nM (pChEMBL 8.079999923706055)
Descriptors: MW 484.56, ALogP 2.02, HBA 7, HBD 3, PSA 139.04, QED 0.61
Compound (SMILES): CN(C(=O)c1ccc(C2CC2)c(OCC2CCCO2)n1)C(C)(C)C
InChIKey: VOJUPIFFPNJQEP-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 332.44, ALogP 3.39, HBA 4, HBD 0, PSA 51.66, QED 0.83
Compound (SMILES): O=c1cc(N[C@H]2CCN(S(=O)(=O)C(F)(F)F)C2)c2cc(C(c3ccc(Cl)cc3)c3nccs3)ccc2[nH]1
InChIKey: KWNLQHPAAMOAJZ-LBOXEOMUSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 > 3299.89 nM (pChEMBL 5.48199987411499)
Descriptors: MW 569.03, ALogP 5.15, HBA 6, HBD 2, PSA 95.16, QED 0.34
Compound (SMILES): Cc1cc(NC(=O)N2CC=C(c3ncc(-c4nnc(CO)o4)cc3Cl)CC2)ccc1OC(F)(F)F
InChIKey: VUYOBJORBPPYGH-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8NER1
Assay: Capsaicin-Based Assay: Two days prior to performing this assay, cells are seeded in poly-D-lysine-coated 96-well clear-bottom black plates (50,000 cells/well) in growth media containing 5 uM PonA (commercially available from Invitrogen) to induce expression of TRPV1. On the day of the assay, the plates are washed with 0.2 mL 1x Hank's Balanced Salt Solution (commercially available from Life Technologies) containing 1 mM CaCl2 and 20 mM HEPES, pH 7.4, and cells are loaded using 0.1 mL of wash buffer containing Fluo-4 (3 uM final). After one hour, the cells are washed twice with 0.2 mL of wash buffer and resuspended in 0.1 mL of wash buffer. The plates are transferred to a FLIPR for assay. 50 uL of test compound diluted with assay buffer (1x Hank's Balanced Salt Solution containing 1 mM CaCl2 and 20 mM HEPES, pH 7.4) are added to the cell plates and incubated for 2 min. The final concentration of the compound is adjusted to range from about 50 picoM to about 3 uM.
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 509.87, ALogP 4.81, HBA 7, HBD 2, PSA 113.61, QED 0.51
Compound (SMILES): NC(=O)c1c(N)ncnc1N1CCC(c2nc(-c3ccc(F)c(C(F)(F)F)c3)cn2CCNCC2CC2)CC1
InChIKey: PNRZJJLWTXOCFE-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 = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 546.57, ALogP 3.56, HBA 8, HBD 3, PSA 127.98, QED 0.28
Compound (SMILES): C[C@@H](Nc1ncnc2[nH]c(-c3ccc(OCC(=O)NO)cc3)cc12)c1ccccc1
InChIKey: RRWPCUPACFSZEW-CQSZACIVSA-N
Target: Receptor tyrosine-protein kinase erbB-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P04626
Assay: Inhibition of N-terminal GST-tagged human HER2 (Lys676 to Val1255 residues) expressed in baculovirus expression system using biotinylated FLT3 at Tyr589 as substrate preincubated for 5 mins followed by substrate addition measured after 30 mins by DELFIA method
Measurement: IC50 <= 100 nM (pChEMBL 7)
Descriptors: MW 403.44, ALogP 3.68, HBA 6, HBD 4, PSA 112.16, QED 0.28
Compound (SMILES): O=[N+]([O-])c1ccc2nc(NCCc3cc[n+](Cc4ccc(F)cc4)cc3)sc2c1
InChIKey: PRXIEXWZIFEMSH-UHFFFAOYSA-N
Target: Acetylcholinesterase (SINGLE PROTEIN), Homo sapiens
UniProt: P22303
Assay: AChE/BuChE Inhibition Assay: The cholinesterase assays were performed using colorimetric method reported by Ellman. The appearance of product was monitored at 412 nm for 5 min using a spectrophotometer. The reaction rates were compared and the percent inhibition due to the presence of test compounds was calculated. Inhibition curve was obtained for each compound. The linear regression parameters were determined and the IC50 values extrapolated.
Measurement: IC50 = 96 nM (pChEMBL 7.019999980926514)
Descriptors: MW 409.47, ALogP 4.33, HBA 5, HBD 1, PSA 71.94, QED 0.28
Compound (SMILES): OC1CCc2c(-c3ccc(C(F)(F)F)c(F)c3)cncc21
InChIKey: XESQDDURLUPHGC-UHFFFAOYSA-N
Target: Cytochrome P450 11B2, mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: P19099
Assay: Cellular Enzyme Assay: 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 ug/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 uM 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 = 5.2 nM (pChEMBL 8.279999732971191)
Descriptors: MW 297.25, ALogP 3.89, HBA 2, HBD 1, PSA 33.12, QED 0.81
Compound (SMILES): CC(C)N1C(=O)C2(N=C1N)c1cc(Br)ccc1C[C@]21CC[C@@H](OC(F)F)CC1
InChIKey: CDVODTJCICQRJM-LPEZOHBNSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: BACE Assay: For each compound being tested, the BACE activity was monitored in a fluorescence quenching assay (FRET) using the ectodomain of BACE (aa 1-454) fused to a myc-his tag and secreted from HEK293/BACEect. cells into OptiMEM (Invitrogen) as enzyme source and a substrate peptide derived from the APP-Swedish mutation which possesses a Cy3-fluorophore at the N-terminus and a Cy5Q-quencher at the C-terminus (Cy3-SEVNLDAEFK-Cy5Q-NH2; Amersham). The substrate was dissolved at 1 mg/mL in DMSO.The assay was performed in the presence of 5 μl OptiMEM (supernatant collected over 24 hours and cleared from cellular debris by centrifugation) containing the ectodomain of BACE, 25 μl water containing the desired concentration of test compound and 1% DMSO, 1 μM substrate peptide, 20 mM NaOAc, pH 4.4 and 0.04% Triton-X100 in a total assay volume of 50 μl in a 384 well plate. In general, 25 μl of compound dilution were given to the plate followed by the addition of 10 μl of BACE containing OptiMEM diluted 1:2 in water with 0.2% Triton X-100. The reaction was started with the addition of 15 μl substrate in NaOAc buffer. The reaction was incubated at 30° C. in a fluorimeter and the cleavage of the substrate was recorded as kinetic for 60 min. at ex: 530 nm, em: 590 nm. Blank wells containing either no inhibitor or no enzyme were included on each plate.The intensity of fluorescence was regressed against time in order to derive velocities of reaction in all 384 wells. These velocities were used for calculating percent inhibition using an uninhibited control containing 1% DMSO and a fully inhibited control incubations performed in the absence of enzyme. IC50 values were calculated by fitting percent inhibition vs. inhibitor concentration using standard software like GraphPadPrism.
Measurement: IC50 = 44.9 nM (pChEMBL 7.349999904632568)
Descriptors: MW 456.33, ALogP 3.93, HBA 4, HBD 1, PSA 67.92, QED 0.75
Compound (SMILES): C[C@H]1Cc2c(nnn2-c2cc(F)ccn2)CN1C(=O)c1cccc(Cl)c1Cl
InChIKey: HTGDPGZNWXWJGU-JTQLQIEISA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: FLIPR Assay: 1321 N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 ul volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250Ã¿ the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 uL of the compound into 300 uL of assay buffer. A further 3x dilution occurred when transferring 50 uL/well of the compound plate to 100 uL/well in the cell plate. Cells were incubateÂ  with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 uL/well of BzATP (final concentration is 250 uM BzATP (human and rat) or 600 (mouse)). The fluorescence change was measured 180 seconds after adding the agonist.
Measurement: IC50 = 390.8 nM (pChEMBL 6.409999847412109)
Descriptors: MW 406.25, ALogP 3.7, HBA 5, HBD 0, PSA 63.91, QED 0.65
Compound (SMILES): Cc1c([C@H]2CN3CCN(C(=O)Cc4ccc(-n5cnnn5)nn4)C[C@H]3CO2)ccc(F)c1C#N
InChIKey: YPAKREDNTQZBQZ-OXJNMPFZSA-N
Target: Voltage-gated inwardly rectifying potassium channel KCNH2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q12809
Assay: Inhibition of human ERG by electrophysiology method
Measurement: IC50 = 33700 nM (pChEMBL 4.46999979019165)
Descriptors: MW 463.48, ALogP 0.6, HBA 10, HBD 0, PSA 125.95, QED 0.55
Compound (SMILES): O=S(=O)(NCC1CC1)c1ccc(Nc2cccnc2Cl)cc1C(F)(F)F
InChIKey: PAKBVBIXBHXATQ-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: Binding Assay: The binding assays were performed by testing the ability of the new compounds to displace the radiolabeled synthetic non-selective cannabinoid agonist [3H]CP55940 (168 Ci/mmol; PerkinElmer) from the human CB1 (hCB1) or human CB2 (hCB2) receptors on membranes derived from stably transfected HEK-293 cells (PerkinElmer). Membranes were diluted in assay buffer (50 mM Tris-HCl, 1 mM EDTA, 1 mM MgCl2, 1 mg/ml BSA, pH=7.4). The amount of membrane was determined for each batch of membranes according to protein binding assay. The minimum amount of membrane that gave 50% specific binding was used for the binding assay. In most assays, binding was tested using 8 μg and 4 μg protein of hCB1 and hCB2 membranes, respectively. The tested compounds were dissolved in DMSO and diluted in assay buffer to a final concentration of 0.1% solvent. Total binding of [3H]CP55940 was evaluated with 1.5 nM to hCB1 and with 0.5 nM to hCB2, according to Kd affinity of [3H]CP55940 for the respective membranes. The ability of the tested compounds to displace [3H]CP55940 was evaluated first at a single concentration point of 100 nM for binding toward hCB2 or hCB1. In certain cases, the displacement was tested at compound concentrations ranging from 0.03 nM to 10 μM. Non-specific binding was measured by the addition of 1 μM of unlabelled CP55940 to the tubes. Binding assays were performed in triplicate in a total volume of 500 μl for 60 minutes at 30° C., in a shaking bath. Free and bound radioligands were separated by rapid filtration through GF/C filter plates (PerkinElmer) that had been presoaked with 0.1% Polyethylenimine (Sigma). Filters were shaken for 1 hour in 7 ml scintillation fluid (PerkinElmer) and radioactivity was determined by liquid scintillation counter (Wallac; PerkinElmer).
Measurement: IC50 = 95.6 nM (pChEMBL 7.019999980926514)
Descriptors: MW 405.83, ALogP 4.19, HBA 4, HBD 2, PSA 71.09, QED 0.71
Compound (SMILES): Cc1cc(-c2cc(OC(F)(F)F)ccc2F)ccc1OCCCC(=O)O
InChIKey: NIWINBARFFCWGP-UHFFFAOYSA-N
Target: Free fatty acid receptor 4 (SINGLE PROTEIN), Mus musculus
UniProt: Q7TMA4
Assay: Agonist activity at mouse GPR120 expressed in CHOK1 cells measured after 60 mins by IP1-HTRF assay
Measurement: EC50 = 79 nM (pChEMBL 7.099999904632568)
Descriptors: MW 372.31, ALogP 4.94, HBA 3, HBD 1, PSA 55.76, QED 0.55
Compound (SMILES): Cn1cc(Nc2nccc(-c3cnc(N4CCC[C@H]4C#N)c(C#N)c3)n2)cn1
InChIKey: KGBHFGYIQUOGQH-INIZCTEOSA-N
Target: Serine/threonine-protein kinase TBK1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UHD2
Assay: Time-Resolved Fluorescence-based Lanthascreen Assay: Kinase inhibition assays (10 μL) were performed at 20° C. in 384-well plate format. Compound IC50 values were determined at the apparent Km for ATP (20 μM) based on a radiometric assay (Invitrogen) using 8 or 10 point curves in duplicate. The final reaction conditions contained 400 nM fluorescein-IkBα substrate (DRHDSGLDSMKDE), 20 μM ATP, 2 nM or 8 nM IKKε or TBK1 kinase respectively, and 3% DMSO in kinase assay buffer consisting of 50 mM HEPES (pH 7.5), 10 mM MgCl, 1 mM EGTA, 0.01% Brij-35.Compound dilutions were prepared from 10 mM DMSO stocks by dilution into DMSO. Compound dilution series were further diluted in kinase assay buffer to give a 12% DMSO stock, the final concentration in the assay being 3% DMSO.The kinase phosphorylation assay was initiated by the addition of the kinase and the reaction was allowed to proceed for 1 h or 2.5 h for IKKε and TBK-1 kinase respectively.
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 371.41, ALogP 2.38, HBA 9, HBD 1, PSA 119.34, QED 0.74
Compound (SMILES): CN(CCCn1c(=O)oc2cc(CNC[C@H](O)c3ccc(O)c4[nH]c(=O)ccc34)ccc21)[C@H]1CC[C@H](OC(=O)C(O)(c2cccs2)c2cccs2)CC1
InChIKey: OPRQRGHVAAWHMO-LNFMTQTQSA-N
Target: Muscarinic acetylcholine receptor M3 (SINGLE PROTEIN), Homo sapiens
UniProt: P20309
Assay: Muscarinic Receptor Binding Assay: The study of binding to human muscarinic M1, M2, M3, M4 and M5 receptors was performed using commercial membranes (Perkin Elmer) prepared from CHO-K1 cells. Radioligand binding experiments were conducted in 96 polypropylene well plates in a total volume of 200 ul. All reagents were dissolved in assay binding buffer (PBS with calcium and magnesium, SIGMA), except compounds that were dissolved in DMSO 100%. Non-specific binding (NSB) was measured in the presence of 1 uM atropine. [3H]-NMS was used as the radioligand at a concentration of 1 nM for M2, M3 and M5 and 0.3 nM for M1 and M4. [3H]-NMS and antagonists were incubated with membranes that express human muscarinic receptors M1, M2, M3, M4 and M5 at concentrations of 8.1, 10, 4.9, 4.5 and 4.9 ug/well, respectively. After an incubation period of two hours with gentle shaking, 150 ul of the reaction mix were transferred to 96 GF/C filter plates (Millipore), previously treated with wash buffer.
Measurement: IC50 = 0.3 nM (pChEMBL 9.520000457763672)
Descriptors: MW 758.92, ALogP 5.2, HBA 13, HBD 5, PSA 170.26, QED 0.09
Compound (SMILES): CN1Cc2c(Cl)cc(Cl)cc2[C@H](c2cccc(S(=O)(=O)NCCOCCOCCNC(=O)Nc3ccc(NC(=O)NCCOCCOCCNS(=O)(=O)c4cccc([C@@H]5CN(C)Cc6c(Cl)cc(Cl)cc65)c4)cc3)c2)C1.Cl.Cl
InChIKey: KBWZTUSFIDBIBN-PNBTXOOZSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P26433
Assay: Inhibition of rat NHE3-mediated sodium-dependant hydrogen ion antiport in Opossum kidney cells preincubated for 60 mins in NaCl-HEPES buffer followed by compound washout with NH4Cl-HEPES buffer containing BCECF-AM dye and subsequent washout with Na+ free HEPES buffer by fluorescence analysis
Measurement: IC50 = 7.943 nM (pChEMBL 8.100000381469727)
Descriptors: MW 1165.06
Compound (SMILES): CC(=O)c1c(C)c2cnc(Nc3ccc(N4CCNCC4)cc3)nc2n(C2CCCC2)c1=O.Cl
InChIKey: UEPHCUWNFIIJRQ-UHFFFAOYSA-N
Target: CDK2/Cyclin A2 (PROTEIN COMPLEX), Homo sapiens
UniProt: P20248
Assay: Inhibition of Cdk2/Cyclin A (unknown origin) using retinoblastoma protein substrate and [gamma32P]ATP incubated for 15 mins by beta plate counter based assay
Measurement: IC50 = 233 nM (pChEMBL 6.630000114440918)
Descriptors: MW 446.56, ALogP 3.57, HBA 8, HBD 2, PSA 92.15, QED 0.58
Compound (SMILES): O=C(OC(C(F)(F)F)C(F)(F)F)N1CCC(C(O)(c2ccc3c(c2)OCO3)c2ccc3c(c2)OCO3)CC1
InChIKey: SXHQLPHDBLTFPM-UHFFFAOYSA-N
Target: Monoglyceride lipase (SINGLE PROTEIN), Homo sapiens
UniProt: Q99685
Assay: Inhibition of human recombinant MAGL using 4-NPA as substrate after 30 mins
Measurement: IC50 = 3 nM (pChEMBL 8.520000457763672)
Descriptors: MW 549.42, ALogP 4.72, HBA 7, HBD 1, PSA 86.69, QED 0.55
Compound (SMILES): CC1(C)CCC(CN2CCN(c3ccc(C(=O)NS(=O)(=O)c4ccc(NCCCN5CCOCC5)c(S(=O)(=O)C(F)(F)F)c4)c(Oc4cccc5[nH]ccc45)c3)CC2)=C(c2ccc(Cl)cc2)C1
InChIKey: ORVZVQHRPILFPL-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The inhibition constant (Ki) is the dissociation constant of an enzyme-inhibitor complex or a protein/small molecule complex, wherein the small molecule is inhibiting binding of one protein to another protein or peptide. Where the Ki for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-XL) is greater than the limits of detection of the assay used. Where the binding selectivity ratio for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the selectivity of a particular compound for Bcl-2 over Bcl-XL is at least as great as the number indicated. Where the Ki for a compound is represented as < (less than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-2) is lower than the limit of detection of the assay used. Inhibition constants were determined using Wang's equation (Wang Z-X).
Measurement: Ki = 24.23 nM (pChEMBL 7.619999885559082)
Descriptors: MW 983.58, ALogP 8.95, HBA 11, HBD 3, PSA 153.38, QED 0.09
Compound (SMILES): CN1Cc2c(Cl)cc(Cl)cc2C(c2cccc(S(=O)(=O)NCCOCCOCCNC(=O)c3ccc(C(=O)NCCOCCOCCNS(=O)(=O)c4cccc(C5CN(C)Cc6c(Cl)cc(Cl)cc65)c4)cc3)c2)C1
InChIKey: NHIXKUDRYQFGKB-UHFFFAOYSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P26433
Assay: Inhibition of rat NHE3 expressed in opossum kidney cells assessed as reduction in Na-HEPES buffer-mediated pH recovery in presence of NEH1 inhibitor ethyl isopropyl amiloride by BCECF-AM dye based fluorescence assay
Measurement: IC50 = 6309.57 nM (pChEMBL 5.199999809265137)
Descriptors: MW 1135.03
Compound (SMILES): COC(=O)Nc1nc2cc(C3(O)c4ccccc4C(=O)N3C(C)c3ccccc3F)ccc2[nH]1
InChIKey: YKIGCHLVTYJFLY-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 = 1000 nM (pChEMBL 6)
Descriptors: MW 460.47, ALogP 4.29, HBA 5, HBD 3, PSA 107.55, QED 0.42
Compound (SMILES): COc1cc(OCc2ccccc2)ccc1/C=C/C(=O)CC(=O)/C=C/c1ccc2cc[nH]c2c1
InChIKey: ZAPPNMGPHQBOBC-SDCOAONHSA-N
Target: Microtubule-associated protein tau (SINGLE PROTEIN), Homo sapiens
UniProt: P10636
Assay: Inhibition of recombinant human TAU three-repeat microtubule-binding domain aggregation expressed in Escherichia coli after 16 hrs by thioflavin T fluorescence assay
Measurement: IC50 = 610 nM (pChEMBL 6.210000038146973)
Descriptors: MW 451.52, ALogP 6.01, HBA 4, HBD 1, PSA 68.39, QED 0.24
Compound (SMILES): CCCCN(CCCC)C(=O)c1nn(-c2ccc(C(=O)NS(=O)(=O)c3ccc4cccc(Cl)c4c3)cc2C(=O)N2CCc3ccccc3C2)c(C)c1Cl
InChIKey: QNJUPSQRBSPXOT-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q07817
Assay: Fluorescence Resonance Energy Transfer Assay (FRET): The assays were performed in black flat-bottom 384-well plates. The final assay volume was 55 μl prepared from additions of Biotin-Bcl-xL (Bcl-xL: GENBANK® Accession No. Q07817), fluoresceinated 18-mer BIM peptide (NH2-YYANFEDGIRRLEQAIWI-[FAM]) (SEQ ID NO: 1), Streptavidin-Terbium FRET detection reagent, and test compounds in assay buffer consisting of 20 mM Sodium Phosphate, 1 mM EDTA, 50 mM NaCl, and 0.05% Pluronic Acid. The reaction was incubated at room temperature for 60 minutes. After 60 minutes, 5 μL of Streptavidin-Terbium FRET detection reagent (Perkin Elmer) was added to the reaction mixture and incubated at room temperature in the dark for 30 mins. The FRET signal generated by the reaction was detected on the Envision Plate Reader Inhibition data were calculated from FRET values generated by the no protein control reactions for 100% inhibition and vehicle-only reactions for 0% inhibition. The final concentration of reagents in the assay was 10 nM Bcl-xL, 5 nM fluo
Measurement: IC50 = 70 nM (pChEMBL 7.159999847412109)
Descriptors: MW 774.77, ALogP 8, HBA 7, HBD 1, PSA 121.68, QED 0.14
Compound (SMILES): COc1cnc(C(=O)Nc2cc(F)c(F)c([C@]3(C)C[C@@H](C(F)(F)F)OC(N)=N3)c2)c(C)n1
InChIKey: JZWAAFAQFIIWCU-SGTLLEGYSA-N
Target: Beta-secretase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P56817
Assay: In Vitro BACE Cell-Based Assay: The cell-based assay measures inhibition or reduction of Aβ40 in conditioned medium of test compound treated cells expressing amyloid precursor protein. Cells stably expressing Amyloid Precursor Protein (APP) were plated at a density of 40K cells/well in 96 well plates (Costar). The cells were cultivated for 24 hours at 37° C. and 5% CO2 in DMEM supplemented with 10% FBS. The test compounds were then added to cells in 10-point dose response concentrations with the starting concentration being either 100 uM or 10 uM. The compounds were diluted from stock solutions in DMSO and the final DMSO concentration of the test compounds on cells was 0.1%. After 24 h of incubation with the test compounds the supernatant conditioned media was collected and the Aβ 40 levels were determined using a sandwich ELISA.
Measurement: IC50 = 85 nM (pChEMBL 7.070000171661377)
Descriptors: MW 459.38, ALogP 3.21, HBA 7, HBD 2, PSA 111.72, QED 0.68
Compound (SMILES): CCOc1ccc(-c2oc3cc(N(C)S(C)(=O)=O)c(-c4ccc5c(n4)-c4cc6c(F)cccc6n4CO5)cc3c2C(=O)NC)cn1
InChIKey: OCVUSFSYTHVTFK-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1b (isolate Japanese) (HCV)
UniProt: P26662
Assay: Inhbition Assay: To initiate an assay, replicon cells were plated in the presence of a dilution series of test compound in the absence of G418. Typically, the assays were performed in a 96-well plate formate for manual operation, or a 384-well plate format for automated assay. Replicon cells and compound were incubated for 96 hours. At the end of the assay, cells were washed free of media and compound, and the cells were then lysed. RNA was quantified indirectly through detection of replicon-encoded NS3/4A protein levels, through an ELISA-based assay with an antibody specific for NS3/4A.
Measurement: IC50 = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 641.68, ALogP 5.82, HBA 9, HBD 1, PSA 128.79, QED 0.23
Compound (SMILES): Cc1c(F)cccc1NC(=O)Nc1ccc(Cl)c(S(=O)(=O)C2CNC2)c1O
InChIKey: JQJOUUXCZZNDSH-UHFFFAOYSA-N
Target: C-X-C chemokine receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P25025
Assay: Displacement of [125I]IL8 from recombinant human CXCR2 expressed in CHO cell membranes measured after 1 hr by TopCount scintillation counting method
Measurement: IC50 < 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 413.86, ALogP 2.88, HBA 5, HBD 4, PSA 107.53, QED 0.58
Compound (SMILES): CC1(C)CC2=NNC(=O)c3c(F)ccc4[nH]c(c2c34)C1
InChIKey: XDQDKCGQZXCZGC-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGN5
Assay: PARP-2 and PARP-3 Enzymatic Assay: PARP-2 and PARP-3 enzymatic assays were conducted using commercial PARP-2/PARP-3 Chemiluminescent Assay Kit (BPS Biosciences) and the protocols with the kits. Briefly, histones were coated in a high binding plate first, and incubated with PARP-2 or PARP-3, and increasing concentrations of compounds for 0.5 h. Then, biotinylated NAD and activated DNA were added to the wells. The biotinylated PARylation product was measured by adding streptavidin-HRP and HRP substrates which produce chemiluminescence.
Measurement: IC50 = 0.7 nM (pChEMBL 9.149999618530273)
Descriptors: MW 271.3, ALogP 2.73, HBA 2, HBD 2, PSA 57.25, QED 0.76
Compound (SMILES): COc1cc(N2CCOCC2)ccc1Nc1nc2c(c(Nc3ccccc3S(=O)(=O)NC3CC3)n1)C1(CC1)CN2
InChIKey: CFPSVRXYURHCRZ-UHFFFAOYSA-N
Target: Protein-tyrosine kinase 2-beta (SINGLE PROTEIN), Homo sapiens
UniProt: Q14289
Assay: Biochemical Assay: To measure inhibitory activity of FAK/PYK2 inhibitors, compounds were first prepare as a 1 mM stock in 100% DMSO and 3-fold serial dilution was performed in a 96-well plate (Corning, 3897) to generate 12 different concentration of 100× stock. A 5 μl of 100× stock of each concentration was added to wells containing 95 μl of 1× reaction buffer (40 mM Tris, pH7.5, mM MgCl2, 1 mM DTT and 1 mM CHAPS) to generate 5× stock. Then 2 μl of 5× stock of each concentration was added to a 384 wells-OptiPlate (PerkinElmer, 6007299). For FAK, 4 μl of 2.5 nM FAK, 1 μl of 8 μg/μl BSA and 1.5 μl of 666 nM ULight-poly Glu, Ala, Tyr (1:1:1) peptide substrate, prepared in above reaction buffer, were added to each well. The reaction was initiated by adding 1.5 μl of 33.3 μM ATP. The reaction was allowed to proceed for 120 min before being quenched with 5 μl of 40 mM EDTA stop buffer prepared in 1×LANCE detection buffer (PerkinElmer, CR97-100). For PYK2, 5 μl of 2 nM PYK2 and 1.5 μl of 666 nM ULight-poly Glu, Ala, Tyr (1:1:1) peptide substrate, prepared in above reaction buffer, were added to each well. The reaction was initiated by adding 1.5 μl of 46.6 μM ATP. The reaction was allowed to proceed for 40 min before being quenched with 5 μl of 40 mM EDTA stop buffer prepared in 1×LANCE detection buffer (PerkinElmer, CR97-100). Upon quenching of the reaction, 5 μl of 8 nM anti-phosphotyrosine antibody was added to each well and incubated for 60 minutes.
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 563.68, ALogP 3.71, HBA 10, HBD 4, PSA 129.74, QED 0.31
Compound (SMILES): CC#Cc1ccnc(-c2ccc3c(c2)C2(N=C(C)C(N)=N2)[C@]2(CC[C@H](OC)CC2)C3)c1
InChIKey: BHCDFMZEDWIROP-AHQGXOPPSA-N
Target: Amyloid-beta precursor protein (SINGLE PROTEIN), Homo sapiens
UniProt: P05067
Assay: sAPPbeta Release Assay: SH-SY5Y cells were cultured in DMEM/F-12 with Glutamax, 10% FCS and 1% non-essential amino acids and cryopreserved and stored at −140° C. at a concentration of 7.5-9.5×106 cells per vial. Thaw cells and seed at a conc. of around 10000 cells/well in DMEM/F-12 with Glutamax, 10% FCS and 1% non-essential amino acids to a 384-well tissue culture treated plate, 100 μL cell susp/well. The cell plates were then incubated for 7-24 h at 37° C., 5% CO2. The cell medium was removed, followed by addition of 30 μL compound diluted in DMEM/F-12 with Glutamax, 10% FCS, 1% non-essential amino acids and 1% PeSt to a final conc. of 1% DMSO. The compounds were incubated with the cells for 17 h (overnight) at 37° C., 5% CO2. Meso Scale Discovery (MSD) plates were used for the detection of sAPPβ release. MSD sAPPβ plates were blocked in 1% BSA in Tris wash buffer (40 μL/well) for 1 h on shake at r.t. and washed 1 time in Tris wash buffer (40 μL/well). 20 μL of medium was transferred to the pre-blocked and washed MSD sAPPβ microplates, and the cell plates were further used in an ATP assay to measure cytotoxicity. The MSD plates were incubated with shaking at r.t. for 2 h and the media discarded. 10 μL detection antibody was added (1 nM) per well followed by incubation with shaking at r.t. for 2 h and then discarded. 40 μL Read Buffer was added per well and the plates were read in a SECTOR Imager.
Measurement: IC50 = 0.78 nM (pChEMBL 9.109999656677246)
Descriptors: MW 412.54, ALogP 4.24, HBA 5, HBD 1, PSA 72.86, QED 0.75
Compound (SMILES): CNC(=O)c1c(-c2ccc(F)cc2)oc2cc(N(C)S(C)(=O)=O)c(-c3ccc4c(n3)-c3c(Cl)c5c(Cl)cccc5n3CO4)cc12
InChIKey: PTFZCZXLDZUYCN-UHFFFAOYSA-N
Target: Genome polyprotein (SINGLE PROTEIN), Hepatitis C virus genotype 1b (isolate Japanese) (HCV)
UniProt: P26662
Assay: ELISA-Based Assay: Measurement of inhibition by compounds was performed using the HCV replicon system. Several different replicons encoding different HCV genotypes or mutations were used. In addition, potency measurements were made using different formats of the replicon assay, including different ways of measurements and different plating formats. See Jan M. Vrolijk et al., A replicons-based bioassay for the measurement of interferons in patients with chronic hepatitis C, 110 J. VIROLOGICAL METHODS 201 (2003); Steven S. Carroll et al., Inhibition of Hepatitis C Virus RNA Replication by 2'-Modified Nucleoside Analogs, 278(14) J. BIOLOGICAL CHEMISTRY 11979 (2003). However, the underlying principles are common to all of these determinations, and are outlined below.Stable neomycin phosphotransferase encoding replicons-harboring cell lines were used, so all cell lines were maintained under G418 selection prior to the assay. Potency was determined using a cell ELISA assay with an antibody to the replicons encoded NS3/4a protease. See Caterina Trozzi et al., In Vitro Selection and Characterization of Hepatitis C Virus Serine Protease Variants Resistant to an Active-Site Peptide Inhibitor, 77(6) J. Virol. 3669 (2003).
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 665.53, ALogP 7.33, HBA 7, HBD 1, PSA 106.67, QED 0.21
Compound (SMILES): CC(C)(C)OC(=O)N1CC=C(c2cc(=O)[nH]c3ccc(C(c4ccc(Cl)cc4)c4ccc(Cl)cc4)cc23)CC1
InChIKey: UBWHVQJTXISRNS-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: cAMP Assay: The following mixtures and buffer solutions were prepared: (a) Buffer 1: HBSS (Mediatech Cat#21-023-CV) with 5 mM HEPES (1 mM stock, Gibco BRL Cat#15630-056) and 0.1% BSA (7.5% stock, Invitrogen Cat#15260-037); (b) Buffer 2: 0.5 mM IBMX (200 mM stock in DMSO, Sigma 15879) in Buffer 1; (c) 1 uM cAMP Standard (50 uM stock, Perkin Elmer Cat# AD0262) diluted in Buffer 2 and serially diluted in Buffer 2, 12 doses at 1/2 dilutions resulting in a dose range of 1 uM to 0.5 nM; (d) d2 labelled cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 6 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); (e) anti-cAMP (CisBio HTRF Detection Kit Cat #62AM4PEB reconstituted with 5 ml dH2O) diluted 1/20 with lysis buffer (CisBio HTRF Detection Kit Cat #62AM4PEB); and (f) Forskolin (Sigma Cat# F6886, 10 mM in DMSO) diluted first in DMSO to 1 mM and then to 1.5 uM in Buffer 2.
Measurement: EC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 561.51, ALogP 8.04, HBA 3, HBD 1, PSA 62.4, QED 0.26