Compounds and methods of identifying, synthesizing, optimizing and profiling protein modulators

A method of testing inhibition or activation by a chemical agent on a selected target protein in a cell, the method comprising:

• a) incubating a first cell which expresses a target protein with a chemical agent, the first cell capable of manifesting a phenoresponse linked to the presence and functional activity of the target protein in the first cell, the phenoresponse also being capable of being modulated by a known modulator;
• b) comparing modulation of the phenoresponse of the first cell treated with the chemical agent to modulation of the phenoresponse of the first cell treated with the known modulator;
• and
• c) determining the chemical agent inhibits or activates the target protein if the modulation of the phenoresponse of the first cell treated with the chemical agent is at least the same degree as the modulation of the phenoresponse of the first cell treated with the known modulator.

A method of testing a chemical agent, comprising:

• a) incubating a first cell with a chemical agent, the first cell expressing a target protein and being capable of manifesting a phenoresponse linked to the presence and functional activity of the target protein in the first cell;
• b) incubating a second cell with the chemical agent, the second cell expressing the target protein at a lower level or not expressing the target protein, the second cell being capable of manifesting a phenoresponse linked to the presence and functional activity of the target protein in the second cell to a lesser extent than the first cell, or not at all;
• c) comparing modulation of the phenoresponse of the first cell treated with the chemical agent to modulation of the phenoresponse of the second cell treated with the chemical agent;
• and
• d) determining the chemical agent inhibits or activates the target protein if the modulation of the phenoresponse of the first cell treated with the chemical agent is greater than the modulation of the phenoresponse of the second cell treated with the chemical agent.

A compound selected as a modulator of a target protein, the compound having improved cellular specificity compared to a known modulator of the target protein, the compound being selected using a method comprising:

• a) measuring modulation of the phenoresponse of a test cell treated with the known modulator, the test cell expressing the target protein at a first level;
• b) measuring modulation of the phenoresponse of a control cell treated with the known modulator, the control cell expressing the target protein at a second level lower than the first level;
• c) measuring modulation of the phenoresponse of the test cell treated with a compound;
• d) measuring modulation of the phenoresponse of the control cell treated with the compound;
• e) determining the cellular specificity gap of the known modulator and the cellular specificity gap of the compound;
• and
• f) identifying the compound as having improved cellular specificity if the cellular specificity gap of the compound is greater than the cellular specificity gap of the known modulator.