Biochemical characterization of desloratadine, a potent antagonist of the human histamine H1 receptor
Introduction
Histamine is a biogenic amine that has natural physiologic functions, including smooth muscle contraction, microvascular permeability, catecholamine release, and neuromodulatory roles in the brain and central nervous system (Hill et al., 1997). Histamine is released upon stimulation of mast cells and is one of the major mediators of the pathophysiology of the allergic response (Slater et al., 1999).
The pharmacology of histamine is mediated through four distinct G-protein coupled receptors classified as H1, H2, H3, and H4. All four human histamine receptors have been cloned, as have bovine and rat histamine H1 receptors DeBacker et al., 1993, Moguilevsky et al., 1994, Gantz et al., 1991, Yamashita et al., 1991, Lovenberg et al., 1999, Oda et al., 2000, Morse et al., 2001. Histamine binding to the histamine H1 receptor results in increases in intracellular calcium ([Ca2+]i) mobilization (Kotlikoff et al., 1987) and is linked to the smooth muscle contraction and edema associated with the allergic response.
Histamine H1 receptor antagonists inhibit the binding of histamine to the histamine H1 receptor, prevent many of histamine's adverse effects, and are considered first-line therapy for the treatment of allergic diseases (Meltzer, 1998). Most of the older histamine H1 receptor antagonists have the potential for adverse central nervous system effects and are characterized by poor receptor specificity. They bind to muscarinic receptors resulting in marked anticholinergic effects (Meltzer, 1998). Newer antihistamines, including loratadine, cetirizine, azelastine, and fexofenadine, have improved side-effect profiles, but some, notably cetirizine and azelastine, can also cause sedation (Slater et al., 1999). In addition, the histamine H1-receptor antagonists, terfenadine and astemizole, demonstrated severe cardiovascular liabilities Woosley et al., 1993, Craft, 1986, including torsades de pointes, a potentially fatal cardiac abnormality characterized by ventricular tachycardia Monahan et al., 1990, Saviuc et al., 1993. This is believed to be due to blockade of the IKr channel, which inhibits the rapid delayed rectifier K+ current and prolonged the action potential (Woosley et al., 1993).
Desloratadine is a new histamine H1 receptor antagonist for the treatment of seasonal allergic rhinitis Handley, 1999, Salmun et al., 2000. Preclinical studies showed desloratadine to be a potent and selective histamine H1-receptor antagonist with relatively low affinities for muscarinic (Ki values of 50, 47, 104 and 320 nM at the human muscarinic M1, M2, M4 and M5 receptors, respectively) and histamine H2 (Ki value of 353 nM) receptors Handley et al., 1997, Kreutner et al., 2000. Desloratadine does not interact with the human ether-a-go-go-related gene (HERG) channel at concentrations up to 10 μM, suggesting a low potential for ventricular arrhythmias. In addition, desloratadine does not cross the blood–brain barrier and does not cause drowsiness or sedation (Hey et al., 2000). Clinical studies have demonstrated that desloratadine effectively improves nasal and nonnasal symptoms of allergic rhinitis and is not associated with adverse cardiovascular or central nervous system effects (Salmun et al., 2000). In this report, we have characterized the affinity, potency, and mechanism of the antagonism of desloratadine in receptor-binding and functional assays at the human histamine H1 receptor cloned from human lung and expressed in Chinese hamster ovary (CHO) cells.
Section snippets
Synthesis of [3H]desloratadine
[3H]Desloratadine was prepared by Tris-triphenylphosphine ruthenium (II) chloride catalyzed exchange with tritiated water (Fig. 1). Desloratadine (20 mg) and Tris-triphenylphosphine ruthenium (II) chloride (2 mg) were suspended in dioxane (100 μl) in a thick-wall glass ampule. Tritiated water (90 at.%, 20 Ci) was distilled in, and the ampule was capped, frozen in liquid nitrogen, evacuated, and flame sealed. The tube was heated for 3 h at 120 °C, after which it was cooled to room temperature
Saturation-binding studies
The affinity of [3H]desloratadine for the human histamine H1 receptor was determined by saturation-binding analysis. The binding of [3H]desloratadine (Kd=1.1±0.2 nM; Bmax=7.9±2 pmol/mg protein) to the human histamine H1 receptor expressed in CHO cells was specific, saturable, and of high affinity (Fig. 2). Scatchard transformation of the [3H]desloratadine saturation isotherm demonstrated that the binding was to a single population of receptor sites (Fig. 2). No specific binding of [3
Discussion
In this report, we have characterized the pharmacology of desloratadine at the human histamine H1 receptor that was cloned from human lung tissue and recombinantly expressed in CHO cells. The human histamine H1 receptor was originally cloned from a human lung cDNA library and expressed in CHO and COS cells. In those studies, [3H]pyrilamine bound with a Kd of 3.7 nM (Moguilevsky et al., 1994) and 1.2 nM (DeBacker et al., 1993). This is the first report characterizing [3H]desloratadine at the
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