Margherita Strolin Benedetti Scientific Advisor, Drug Metabolism and Pharmacokinetics, UCB Pharma S.A., France; , Michel Gillard Head of Cellular and Molecular Biology, UCB Pharma S.A., Belgium; , Rhys Whomsley Head, In Vitro Development ADME Laboratory, UCB Pharma S.A., Belgium; , Martin Church Professor of Immunopharmacology, Southhampton General Hospital

Originally printed in:
» European Respiratory Disease 2007 - Issue II

Receptor Agonism and Antagonism and In Vivo Receptor Occupancy

The affinity of a ligand for a receptor determines its degree of receptor occupancy at a given concentration. However, after binding the ligand will either activate the receptor if it is an agonist or prevent its activation if it is an antagonist. Competitive antagonism is based on the principle that agonist and antagonist binding to the receptor are mutually exclusive and that when both agonist and antagonist are present concomitantly they will compete for receptor binding. The ability of a competitive antagonist to influence the receptor occupancy by an agonist is determined by its affinity for the receptor and by the concentrations of both the agonist and antagonist. The important characteristic of competitive antagonism is that the antagonism can always be reversed by increased concentrations of the agonist. However, for slowly reversible or irreversible antagonism, increasing the agonist concentration will not result in a reversal of the effects of the antagonist.

Histamine H1 Receptors

Histamine, released from storage or producing cells into the extracellular space, acts via G-protein-coupled receptors.¹ There are four major subtypes of histamine receptors – H1 to H4 – which differ in their location, their coupling to second messengers and their affinity for histamine. All subtypes can be stimulated by histamine, whose actions can be antagonised by subtype-specific antagonists.²

The human H1 receptor is a 487-amino acid protein with seven transmembrane domains. It is believed that histamine interacts with amino acids in the third and fifth transmembrane domains. In addition, different histamine H1 receptor agonists and antagonists may also bind to different portions of the receptor complex. Stimulation of H1 receptors leads to the hydrolysis of phosphatidyl 4,5-biphosphate and to the formation of inositol-1,4,5 triphosphate and 1,2-diacylglycerol. Inositol-1,4,5-triphosphate mobilises intracellular calcium, while diacylglycerol activates protein kinase C. In accordance, histamine has been shown to induce the production of inositol phosphates and consequently to increase intracellular calcium. The increase in intracellular calcium explains the wide variety of pharmacological effects of H1-receptor stimulation.³

Histamine H1 receptors are involved in the pathological processes of allergy. Clinical trials of H1 receptor antagonists have demonstrated the efficacy of these agents in reducing the sneezing, pruritus and rhinorrhoea associated with allergic rhinitis.

There are very few reports on human H1-receptor polymorphism. A recent paper by Swan et al.⁴ revealed a polymorphism, but in the promotor region of the gene. This may affect the regulation and the expression level of the receptor, but not its affinity towards ligands. Another article by Hong et al.⁵ reports a polymorphism in the coding region of the receptor located in the third intracellular loop at position 349, where a glutamate is replaced by an aspartate. Since it is a conserved mutation (these are two acidic amino acids) and because of its localisation (outside the transmembrane regions), it is not expected to have an influence on antihistamine affinity.

Histamine Metabolism

The histamine released from storage or producing cells into the extracellular space must be inactivated to terminate its effect via histamine receptors on target cells. To be inactivated, histamine must be removed from the extracellular space and metabolised into inactive metabolites. Histamine is metabolised by two main catabolic pathways: oxidative deamination by diamine oxidase (DAO) to imidazole acetic acid and methylation by histamine N-methyltransferase (HNMT) to telemethylhistamine, which is then oxidatively deaminated to methylimidazole acetic acid by monoamine oxidase.⁶ As histamine is unable to easily enter the intracellular space where the enzymes responsible for its metabolism are located, high-affinity transporters for histamine have been considered to be required, and indeed organic cation transporters (OCT)-2 and -3 appear to be important for histamine transport and/or inactivation.^7,8

Tissue Localisation of H1 Receptors, Concentrations of H1 Antihistamines at the Receptor Site and Volume of Distribution of H1 Antihistamines

The H1 receptor is abundant in human respiratory tissues and in various cells of the immune system that play a role in allergic syndromes. H1 receptors are present on T cells, B cells, monocytes, lymphocytes, endothelial cells (e.g. endothelial cells of nasal mucosal blood vessels), gland cells, smooth-muscle cells (e.g. airway and gastrointestinal smooth muscle), neuronal cells, conjunctival epithelial cells and keratinocytes. This list is not intended to be exhaustive, but shows that H1 receptors are widely distributed.^2,3,9

As for every drug administered orally, the prerequisite in order for an H1 antagonist to interact with the receptors is to have a high absolute bioavailability. This means that most of the intact drug should reach the general circulation and should therefore not only be absorbed, but also not be extensively metabolised by enzymes present in the gastrointestinal tract and in the liver.

However, plasma concentrations do not approximate drug concentrations at the receptor site if the drug is bound to plasma proteins. Classically, free plasma concentrations, calculated from plasma concentrations using plasma protein binding, are used as an approximation of the drug concentration at receptor sites, as only unbound drug is capable of entering and leaving the plasma and tissue compartments. However, this assumes the absence of an active transport mechanism; it is therefore important to know whether an H1 antihistamine is a substrate of transporters and which types of transporters are present in the different tissues in which H1 receptors are located, e.g. skin¹⁰ or brain.¹¹

The drug concentrations at receptor sites can also be estimated by an alternative approach, i.e. by using the volume of distribution of the drug, as described by Strolin Benedetti et al. for levocetirizine and desloratadine.¹² As the histamine H1 receptors are localised in the cellular membrane, an H1 antihistamine does not need to be distributed inside the cells to be effective and a volume of distribution close to that of extracellular water should be optimal to obtain a high level of receptor occupancy. This is in agreement with the small volume of distribution observed for highly active H1 antihistamines.^13,14 Moreover, an H1 antihistamine should not impair histamine metabolism: it should therefore neither be an inhibitor of OCT-2 and OCT-3 nor an inhibitor of HNMT and DAO.

Receptor Affinity of H1 Antihistamine In Vivo

The affinity values reported for antagonists – and for the H1 antihistamines in particular – are generally obtained in vitro in conditions not simulating the physiological or pathological conditions, whereas in reality in vivo the antagonist interacts with its receptors in a microenvironment defined by the physiopathological state of the subject. The affinity of an antagonist for its receptor, although necessarily evaluated in vitro, must therefore be estimated in conditions as close as possible to those present in the target tissue(s) during the illness. For this purpose, parameters to be taken into account in the in vitro experiments are temperature (body temperature), incubation time (allowing drugs with slow binding kinetics to reach equilibrium with the receptor) and a suitable buffer (e.g. containing physiological concentrations of NaCl and allowing a pH value that is close to that present in the target tissue containing the receptors to be achieved). For example, as acidosis appears to be a hallmark of inflammatory processes, the affinity of antagonists to be used in inflammatory conditions should be investigated at the pertinent pH in order to better predict their receptor occupancy. The above-mentioned parameters (e.g. pH) do not necessarily affect the affinity of the different H1 antistamines to the same extent, as has been demonstrated by measuring the affinity of levocetirizine, fexofenadine and desloratadine at different pHs.^15–17

Conclusion

In order to be of some relevance for the pharmacodynamic effect and/or the clinical efficacy of the drug, the estimation of receptor occupancy by an antagonist in general, and an H1 antihistamine in particular, requires knowledge of the drug affinity for the receptors in conditions as close as possible to those encountered in the target pathology (e.g. rhinitis, urticaria, inflammatory processes) and knowledge of the unbound concentrations of the drug in the target tissue containing the receptors (e.g. unbound concentrations in skin).

An orally administered H1 antihistamine will give a high fraction of receptors occupied only if it is well absorbed, is not extensively metabolised in the gastrointestinal tract and the liver, can attain the target tissues/cells containing the H1 receptors at sufficiently high free concentrations and its affinity, evaluated in conditions simulating as much as possible the physiopathological conditions, is sufficiently high.

The estimation of receptor occupancy by an H1 antihistamine, as well as the kinetics of the occupancy, can be of great help in interpreting the data obtained in the pharmacodynamic and/or clinical studies and in the selection/prediction of the active doses and administration schedules of the H1 antihistamines. However, for an exhaustive interpretation of the significance of the percentages of receptor occupancy by a given posology of an H1 antihistamine, it would also be important to know concomitantly the concentration of the agonist (histamine) at the receptor site in the pathophysiological condition examined, particularly if the H1 antihistamine is a competitive antagonist.