Interrogating 7TM receptors: Does texture in the question yield greater texture in the answer?

Abstract

The key to detecting and classifying drug effect at seven transmembrane (7TM) receptors is the pharmacological assay. Drug discovery had been rooted in testing of molecules on intact animal tissue until technology provided high-throughput binding assays for screening. While this allowed for the testing of large numbers of molecules, it also limited detection to molecules that interfere with the interaction of the receptor with a defined probe (i.e., radioligand). The ability to monitor functional changes in cells (recombinant or natural) provided a huge leap forward. Earlier functional assays were tied to specific signaling pathways (i.e., cyclic AMP and calcium) but now label-free assays in live cells provide the opportunity to detect more ligands and more fully characterize their efficacy. These ideas will be discussed in terms of harnessing the phenomenon of “functional selectivity” for therapeutic advantage.

Keywords::

• Receptor theory
• new drug discovery
• GPCR signaling
• pathway analysis

Acknowledgements

Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.

Appendix

A receptor [R] is activated by an agonist [A] (equilibrium dissociation constant K_A) to subsequently interact with G proteins [G₁] or [G₂] or with β-arrestin (denoted [BAR]) with equilibrium dissociation constants K₁, K₂, and K_B, respectively. The β-arrestin-receptor complex also then binds to a MAPKinase (denoted [MAP]) with equilibrium dissociation constant K_BM. The receptor conservation equation is:

where [R_tot] is the total receptor species, [R] unbound receptor, [AR] agonist bound receptor, [ARG₁] and [ARG₂] ternary species of agonist-receptor and either G protein [G₁] or [G₂], [ARB] is the agonist-bound receptor/β-arrestin complex and [ARB-MAP] the activated MAPKinase resulting from receptor activation.

The following equilibrium equations determine the species formed:

From these it can be shown that the amount of [ARG₁] complex (as a fraction of [R_tot] is given by:

Similarly, the total species yielding cellular response (assumed to [ARG₁] + [ARG₂] + [ARB-MAP]) is

These responses are processed through a forcing function to simulate saturable processing of stimulus by the cell similar to the operational model for drug response (57). Thus, the response from each species is given by: