## Endpoint to Predict
- Endpoint scope: assay
- Measurement type: IC50
- Label type: continuous
- Binary semantics: pchembl
- Assay description: Screen Quest Fluo-8 No Wash Calcium Assay Kit: Ca2+ influx was measured in HEK-293 cells stably transfected with the receptor using Screen Quest Fluo-8 No Wash Calcium Assay Kit (AAt Bioquest®). Briefly, once inside the cells, the lipophilic blocking groups of Fluo-8 are cleaved by non-specific cell esterases, resulting in a negatively-charged fluorescent-dye that stays inside cells. Its fluorescence increases upon binding to calcium. When HEK-293/P2X7 cells were stimulated with BzATP, Ca2+ entered the cells and the fluorescence of Fluo-8 NW increaseed. The dye absorption spectrum was compatible with excitation at 488 nm by argon laser sources and its emission wavelength was in the range of 515-575 nm.To routinely test the compounds, HEK-293 cells stably transfected with rat P2X7R were seeded overnight in growth medium at 10000, 15000 or 20000 cells/well in 384-well plate, according to the level of response after thawing. 24 hours later, the medium was removed and the cells were pre-loaded at RT for 1 hour with 20 μL/w of Fluo-8 NW prepared in Tyrode 0.3 mM Ca2+/Mg2+-free. Compounds of the invention were tested at 8 concentrations (4 replicates for each concentration): 10-3.16-1-0.32-0.1-0.03-0.01 and 0 μM, in the same plate.Compounds were tested at FLIPRTETRA according to the following method: first injection at FLIPRTETRA of 10 μL of 3× test compound (in Tyrode's buffer 0.3 mM Ca2+/Mg2+-free+DMSO 0.5% final concentration) 5′ incubation second injection at FLIPRTETRA of 15 □L of 3× BzATP at ECK) (in Tyrode's buffer 0.3 mM Ca2+/Mg2+-free+BSA 0% final concentration) Fluorescence recording for 3′Between one plate and the following, tips were extensively washed with water, then with 100% DMSO and finally with water to avoid carry-over inside the tips.The effect of the test compounds was measured as percent inhibition vs a reference antagonist and IC50 values were calculated accordingly.
- Assay organism: Rattus norvegicus
- Target name: P2X purinoceptor 7
- Target type: SINGLE PROTEIN
- Target organism: Rattus norvegicus

## Reference Molecule A
- Canonical SMILES: O=C(NCC(c1scnc1C(F)F)N1CCC(Oc2ccc(Cl)cc2)CC1)c1c(F)cccc1Cl
- Properties:
- ChEMBL ID: CHEMBL5847968
- Molecular weight: 544.43
- Heavy atoms: 35
- Hydrogen bond donors: 1
- Hydrogen bond acceptors: 5
- TPSA: 54.46
- LogP: 6.54
- Rotatable bonds: 8
- Ring count: 4
- Aromatic rings: 3
- Formal charge: 0
- Fraction Csp3: 0.33
- QED: 0.35

## Known Reference Result on This Endpoint
- Molecule A clean result: median pChEMBL = 5.69
- This result is provided as context for predicting Molecule B on the same endpoint.

## Reference Molecule A Other ChEMBL Evidence
- type IC50; relation =; value 2022; units nM; pChEMBL 5.69; comment 912935

## Query Molecule B
- Canonical SMILES: O=C(NCC(c1scnc1C(F)F)N1CCC(Oc2cc(F)cc(F)c2)CC1)c1cccc2ncccc12
- Properties:
- ChEMBL ID: CHEMBL5849486
- Molecular weight: 544.57
- Heavy atoms: 38
- Hydrogen bond donors: 1
- Hydrogen bond acceptors: 6
- TPSA: 67.35
- LogP: 5.92
- Rotatable bonds: 8
- Ring count: 5
- Aromatic rings: 4
- Formal charge: 0
- Fraction Csp3: 0.3
- QED: 0.28

## Molecule Similarity
- Weighted Tanimoto: 0.62

Answer: 6.67