Gpcr screening method to identify non-hallucinogenic compounds
AssigneeThe Regents of the University of California
InventorsChunyang DONG, Calvin LY, David OLSON, Lin TIAN
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Abstract
New fluorescent biosensors are provided for use in methods of detecting a ligand-induced hallucinogenic conformational change of a G Protein-Coupled Receptor (GPCR), detecting a hallucinogenic compound, detecting a non-hallucinogenic antidepressant compound, measuring the hallucinogenic potential of a compound, measuring the antipsychotic potential of a compound, and suitable kits thereof.
Claims54
A method of detecting a ligand-induced hallucinogenic conformational change of a G Protein-Coupled Receptor (GPCR), the method comprising:
- contacting the ligand with a fluorescent biosensor under conditions for the ligand to bind to the fluorescent biosensor, wherein the fluorescent biosensor comprises the GPCR, and a circularly permuted green fluorescent protein (cpGFP) integrated into the third intracellular loop (IL3) of the GPCR;
- and
- measuring the change in fluorescence of the biosensor, thereby detecting the conformational change.
A method of detecting a hallucinogenic compound, the method comprising:
- contacting a compound with a fluorescent biosensor under conditions for the compound to bind to the fluorescent biosensor, wherein the fluorescent biosensor comprises a G Protein-Coupled Receptor (GPCR), and a circularly permuted green fluorescent protein (cpGFP) integrated into the third intracellular loop (IL3) of the GPCR;
- and
- measuring the change in fluorescence of the biosensor, wherein an increase in fluorescence indicates the presence of the hallucinogenic compound, thereby detecting the hallucinogenic compound.
A method of detecting a non-hallucinogenic antidepressant compound, the method comprising:
- contacting a compound with a fluorescent biosensor under conditions for the compound to bind to the fluorescent biosensor, wherein the fluorescent biosensor comprises a G Protein-Coupled Receptor (GPCR), and a circularly permuted green fluorescent protein (cpGFP) integrated into the third intracellular loop (IL3) of the GPCR;
- and
- measuring the change in fluorescence of the biosensor, wherein a decrease in fluorescence indicates the presence of the non-hallucinogenic antidepressant compound, thereby detecting the non-hallucinogenic antidepressant compound.
A method of identifying a hallucinogenic compound from a non-hallucinogenic compound, the method comprising:
- contacting a compound with a fluorescent biosensor under agonist conditions and measuring a first fluorescence signal of the compound, wherein an increase in the first fluoresence signal compared to a first control indicates the compound is hallucinogenic;
- contacting the compound with the fluorescent biosensor under antagonist conditions and measuring a second fluoresence signal of the compound, wherein a decreased second fluoresence signal compared to a second control indicates the compound is non-hallucinogenic;
- and
- combining the first fluoresence signal and the second fluorescence signal to calculate a ligand score where a positive ligand score identifies the compound as a hallucinogenic compound and a negative ligand score identifies the compound as a non-hallucinogenic compound.
A fluorescent biosensor comprising:
- a 5-HT2A receptor;
- and
- a circularly permuted green fluorescent protein (cpGFP) integrated in the third intracellular loop (IL3) of the 5-HT2A receptor.
Dependencies are inferred from the claim text.