Making Sense of Pharmacology: Inverse Agonism and Functional Selectivity

Abstract

Constitutive receptor activity/inverse agonism and functional selectivity/biased agonism are 2 concepts in contemporary pharmacology that have major implications for the use of drugs in medicine and research as well as for the processes of new drug development. Traditional receptor theory postulated that receptors in a population are quiescent unless activated by a ligand. Within this framework ligands could act as agonists with various degrees of intrinsic efficacy, or as antagonists with zero intrinsic efficacy. We now know that receptors can be active without an activating ligand and thus display "constitutive" activity. As a result, a new class of ligand was discovered that can reduce the constitutive activity of a receptor. These ligands produce the opposite effect of an agonist and are called inverse agonists. The second topic discussed is functional selectivity, also commonly referred to as biased agonism. Traditional receptor theory also posited that intrinsic efficacy is a single drug property independent of the system in which the drug acts. However, we now know that a drug, acting at a single receptor subtype, can have multiple intrinsic efficacies that differ depending on which of the multiple responses coupled to a receptor is measured. Thus, a drug can be simultaneously an agonist, an antagonist, and an inverse agonist acting at the same receptor. This means that drugs have an additional level of selectivity (signaling selectivity or "functional selectivity") beyond the traditional receptor selectivity. Both inverse agonism and functional selectivity need to be considered when drugs are used as medicines or as research tools.

Figure 1.

Simulated concentration-response curves to competitors with different intrinsic efficacies on the response to a full agonist (A) or a full inverse agonist (B). (A) Occupancy of the receptor by the full inverse agonist produces 175 units of response. Competition with a ligand of lower intrinsic efficacy reduces the response of the full agonist such that when occupancy of the receptor has been fully replaced by the competitor, the response remaining is due to the competitor and is dependent on the maximal response produced by the competitor. (B) Occupancy of the receptor by the full inverse agonist reduces the basal response (arbitrarily denoted here as 100 units) to 30 units. As a result of competition produced by ligands with higher intrinsic efficacy, the response of the full inverse agonist is reduced to become commensurate to the efficacy of the competitor. Note, the response elicited by the full inverse agonist is not zero, as there remains constitutive activity of signaling molecules (e.g., G proteins) and effectors in the system capable of producing 30 units of response in the absence of constitutive receptor activity.

Figure 2.

Two-state model of receptor function. In this model, receptors in a population exist in equilibrium between an inactive conformation (R) and an active conformation (R*). The proportion of receptors in the active conformation is defined by an allosteric transition constant (L), which is based on the number and strength on intramolecular stabilizing contacts and thus is dependent on the receptor protein structure. The magnitude of response is dependent on the quantity of active receptors, and the efficiency of receptor-effector coupling (K_e). Ligands (A) have affinity for both R (1/K_A) and R* (1/K_A*). Depending on the relative affinities for R vs R*, a ligand can act as an agonist, and inverse agonist or an antagonist. Ligands with higher affinity for R* than R (K_A/K_A*>1) will enrich the population of active receptors (and deplete the population of inactive receptors), leading to increased response, thereby acting as agonists. Conversely, a ligand with higher affinity for R over R* will enrich the population of receptors in the inactive conformation, depleting the population of active conformation receptors and thereby reducing the ongoing response acting as inverse agonists. The efficacy of agonists and inverse agonists is dependent on how far removed the K_A/K_A* ratio is from unity. Ligands with equal affinity for R and R* will not alter the quantity of active receptors and thus not change the ongoing level of responsiveness. However, the presence of an antagonist that can occupy the receptor population will reduce the likelihood of receptor occupancy by other ligands.

Figure 3.

Effects of antagonists and inverse agonists in systems with or without constitutive receptor activity and endogenous agonist tone. (A) In a system where there is no endogenous agonist action and no constitutive receptor activity, application of inverse agonists or antagonists will not alter the basal level of response. (B) When there is constitutive receptor activity but no endogenous agonist action, an antagonist will not alter the basal level of response. By reducing constitutive receptor activity, an inverse agonist will reduce the basal response. (C) In a system with only endogenous agonist tone (no constitutive receptor activity), both antagonists and inverse agonists will reduce the ongoing agonist-dependent response equally as both will reduce receptor occupancy by the agonist. (D) When there is both constitutive receptor activity and action of an endogenous agonist, an antagonist will reduce the component of the response that is due to the endogenous agonist. An inverse agonist will reduce both the endogenous agonist component, but also will reduce constitutive receptor activity; therefore, the effect of the inverse agonist will be greater than that of the antagonist.

Figure 4.

Drug selectivity. (Top) Receptor selectivity is based on differential affinity for different receptor subtypes. Affinity of drugs A and B is reflected by the thickness of the arrows. As shown, drugs A and B have high affinity for the magenta colored receptor and low affinity for the green colored receptor. (Bottom) Functional selectivity is based on differential efficacy of a drug to regulate the activity of various signaling pathways coupled to a single receptor subtype. Signaling selectivity is illustrated as thickness of the arrows between the drug-activated receptor and the cellular signaling pathway. As shown, the selectivity profile for drug A is ERK>ß-arrestin>PLA₂>PLC, whereas that for drug B is PLC>ß-arrestin>PLA₂>ERK. If PLC signaling led to a therapeutic benefit and/or ERK signaling led to an adverse effect, drug B would be the preferred drug. Abbreviations: ß-arr, ß-arrestin; ERK, extracellular signal-regulated kinase; PLA2, phospholipase A2; PLC, phospholipase C.

Figure 5.

Three-state model of receptor function. The simplest multi-active state model of receptor function is the 3-state model, where receptors in a population can adopt either an inactive conformation (R), or 1 of 2 active conformation (R* and R**). As in the 2-state model described in Figure 2, the active receptor conformations, R* and R** are in equilibrium with the inactive conformation (R), as defined by the allosteric transition constants, L and M. The magnitude of response is dependent on the quantity of receptors in an active conformation and the efficiency of receptor-effector coupling (K_e). Thus, the magnitude of constitutive activity can differ for Response 1 vs Response 2, either because L and M are different or K_e1 and K_e2 differ, or both. Ligands have affinity for all 3 receptor conformational states (K_A, K_A*, and K_A**), and ligand efficacy is dependent on the differential affinity values for the 3 conformations. With this model, it is possible that a ligand with disproportionately high affinity for R* vs R and R** could act as a strong agonist for Response 1 (due to enrichment of the R* population), however act as an inverse agonist for Response 2 due to depletion of R**. Thus the same ligand could be simultaneously both an agonist and an inverse agonist, acting via the same receptor. It is important to keep in mind that this model is a pronounced oversimplification on many levels. It is likely that receptors can adopt many more than 3 conformations. Moreover, although this model depicts Response 1 being controlled by R* and Response 2 controlled by R**, it is certainly possible that each active conformation could regulate both responses with different K_e (e.g., K_e1a and K_e1b) values. Also, the model as presented shows that for R* to transition to the R** conformation, it must first become R. This need not happen as it is possible that R* could directly transition to R**. Although likely oversimplified (e.g., Ke associated with R*(*) need not equal Ke associated with AR*(*)), this model was able to account for the behavior of 5-HT_2C agonists to differentially regulate PLC and PLA₂ signaling (Berg et al., 1998).