Screening for positive allosteric modulators: assessment of modulator concentration-response curves as a screening paradigm.

Targeting allosteric binding sites represents a powerful mechanism for selectively modulating receptor function. The advent of functional assays as the screening method of choice is leading to an increase in the number of allosteric modulators identified. These include positive allosteric modulators that can increase the affinity of the orthosteric agonist and potentiate the evoked response. A common method for screening for positive allosteric modulators is to examine a concentration-response (C/R) curve to the putative modulator in the presence of a single, low concentration of agonist. The study reported here has used data simulations for positive allosteric modulators according to the allosteric ternary complex model to generate modulator C/R curves. The results are then compared to the mechanistic parameters used to simulate the data. It is clear from the simulations that the potency of a positive modulator C/R curve in a screening assay is the product of both its affinity and positive cooperativity. However, it is often difficult to tell which parameter dominates the response; not knowing the actual affinity or cooperativity of a ligand may have consequences for receptor selectivity. Further modeling demonstrates that the use and choice of single agonist concentration, as well as changes in the agonist curve Hill slope, can have significant effects on the modulator C/R curve. Finally, the quantitative relationship between modulator C/R curves and the allosteric ternary complex model is explored. These simulations emphasize the importance of careful interpretation of screening data and of conducting full mechanism of action studies for positive allosteric modulators.