## Endpoint to Predict
- Endpoint scope: assay
- Measurement type: Ki
- Label type: continuous
- Binary semantics: pchembl
- Assay description: TR-FRET Binding Assay: A Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) assay that measures the displacement of the 10mer-Thr-FAM probe in response to compound treatment was performed for compounds wherein the IC50 from FPA assay using 10mer-Thr-FAM was below the lower assay IC50 limit 1 nM. Excess 10mer-Thr-FAM probe was utilized with His-tagged WDR5 in conjunction with a commercial anti-His antibody containing a Terbium label. The LanthaScreen Elite Tb-anti-HIS Antibody from ThermoFisher Scientific was used for this purpose. This Tb-anti-HIS has an excitation/emission of 340 nm and 490 nm, respectively. The 10mer-Thr-FAM probe when bound to WDR5 will undergo a FRET interaction with the Tb-anti-HIS and emit at 520 nm. The ratio of the 520 and 495 signals are then utilized to generate a dose-response curve to calculate an IC50 value. By virtue of FRET there is little to no background fluorescence interference from 10mer-Thr-FAM probe allowing an excess of the probe to be used permitting an increase in the lower limit of the calculated Ki when testing against highly potent inhibitors with Ki<<1 nM. WDR5-His Tag (423, residues 24-334) is expressed and purified in our lab in sufficient quantities for screening. 10mer-Thr-FAM peptide is used anywhere from 15 to 150 nM depending on the window of sensitivity required. WDR5-His tag protein is used at 2 nM. A source plate is prepared using an Echo Liquid Handler, which distributes the compounds to the assay plate (white, flat-bottom; OptiPlate) in a 10-point, 3-fold dilution schemes with a top concentration of either 5 or 20 μM depending on the anticipated potency of the compounds, in a final volume of 20 μL. A final target (WDR5)/Tb-Ab concentration of 2 nM/1 nM is dispensed from appropriate stock solutions, respectively. The final DMSO concentration in each well of the assay plate is 1% or lower. As before the plate is covered, shielded from light, and incubated for 60 minutes at room temperature with rocking. Anisotropy is then measured on a Biotek Cytation 3 at excitation wavelength of 340 nm, and emission wavelengths of 495 nm and 520 nm. Working buffer conditions (pH 7) are similar to that in FPA above. TR-FRET signal is plotted and IC50 and Ki values are calculated in the same manner as the fluorescence polarization anisotropy based competition assays.
- Target name: WD repeat-containing protein 5
- Target type: SINGLE PROTEIN
- Target organism: Homo sapiens

## Reference Molecule A
- Canonical SMILES: Cc1nc(F)ccc1-c1cc(Cn2ccnc2)cc(C(=O)NC(C2CC2)C2CC2)c1
- Properties:
- ChEMBL ID: CHEMBL5873530
- Molecular weight: 404.49
- Heavy atoms: 30
- Hydrogen bond donors: 1
- Hydrogen bond acceptors: 3
- TPSA: 59.81
- LogP: 4.36
- Rotatable bonds: 7
- Ring count: 5
- Aromatic rings: 3
- Formal charge: 0
- Fraction Csp3: 0.38
- QED: 0.6

## Known Reference Result on This Endpoint
- Molecule A clean result: median pChEMBL = 8.34
- This result is provided as context for predicting Molecule B on the same endpoint.

## Reference Molecule A Other ChEMBL Evidence
- type Ki; relation =; value 4.6; units nM; pChEMBL 8.34; comment 968712

## Query Molecule B
- Canonical SMILES: COc1cc(CN(C)C(=O)c2cc(CNC3=NCCN3)cc(-c3ccc(F)nc3C)c2)cc(OC)c1
- Properties:
- ChEMBL ID: CHEMBL5875167
- Molecular weight: 491.57
- Heavy atoms: 36
- Hydrogen bond donors: 2
- Hydrogen bond acceptors: 7
- TPSA: 88.08
- LogP: 3.53
- Rotatable bonds: 8
- Ring count: 4
- Aromatic rings: 3
- Formal charge: 0
- Fraction Csp3: 0.3
- QED: 0.47

## Molecule Similarity
- Weighted Tanimoto: 0.35

Answer: 7.92