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Differential Regulation of Thermodynamic Binding Forces of Levocetirizine and (S)-Cetirizine by Lys191 in Human Histamine H₁ Receptors.

ABSTRACT: Cetirizine is a zwitterionic second-generation antihistamine containing R- and S-enantiomers, levocetirizine, and (S)-cetirizine. Levocetirizine is known to have a higher affinity for the histamine H₁ receptors than (S)-cetirizine; ligand-receptor docking simulations have suggested the importance of the formation of a salt bridge (electrostatic interaction) between the carboxylic group of levocetirizine and the Lys191 residue at the fifth transmembrane domain of human histamine H₁ receptors. In this study, we evaluated the roles of Lys191 in the regulation of the thermodynamic binding forces of levocetirizine in comparison with (S)-cetirizine. The binding enthalpy and entropy of these compounds were estimated from the van 't Hoff equation, by using the dissociation constants obtained from their displacement curves against the binding of [³H]mepyramine to the membrane preparations of Chinese hamster ovary cells expressing wild-type human H₁ receptors and their Lys191 mutants to alanine at various temperatures. We found that the higher binding affinity of wild-type H₁ receptors for levocetirizine than (S)-cetirizine was achieved by stronger forces of entropy-dependent hydrophobic binding of levocetirizine. The mutation of Lys191 to alanine reduced the affinities for levocetirizine and (S)-cetirizine, through a reduction in the entropy-dependent hydrophobic binding forces of levocetirizine and the enthalpy-dependent electrostatic binding forces of (S)-cetirizine. These results suggested that Lys191 differentially regulates the binding enthalpy and entropy of these enantiomers, and that Lys191 negatively regulates the enthalpy-dependent electrostatic binding forces of levocetirizine, contrary to the predictions derived from the ligand-receptor docking simulations.

SUBMITTER: Hishinuma S

PROVIDER: S-EPMC6321019 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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Differential Regulation of Thermodynamic Binding Forces of Levocetirizine and (S)-Cetirizine by Lys191 in Human Histamine H₁ Receptors.

Hishinuma Shigeru S Tamura Yuri Y Kobayashi Chihiro C Akatsu Chizuru C Shoji Masaru M

International journal of molecular sciences 20181215 12

Cetirizine is a zwitterionic second-generation antihistamine containing R- and S-enantiomers, levocetirizine, and (S)-cetirizine. Levocetirizine is known to have a higher affinity for the histamine H₁ receptors than (S)-cetirizine; ligand-receptor docking simulations have suggested the importance of the formation of a salt bridge (electrostatic interaction) between the carboxylic group of levocetirizine and the Lys191 residue at the fifth transmembrane domain of human ...[more]

PMID: 30558340

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Project description:ObjectiveLevocetirizine, the R-enantiomer of cetirizine, is classified as a second generation antihistamine used for the treatment of allergic disorders. This study aimed to compare exposure to levocetirizine when given as levocetirizine oral solution (OS) 5 mg to that when given as cetirizine dry syrup (DS) 10 mg, which contains equal proportions of levocetirizine and dextrocetirizine, in healthy Japanese male subjects.MethodsThe study was conducted in an open-label, single dose, randomized and two-way cross-over design. Eligible subjects were allocated to one of two groups and received either levocetirizine OS 5 mg or cetirizine DS 10 mg under fasting conditions, and the alternate treatment after a 7-days washout period. Serial blood samples were taken after each administration, and plasma levocetirizine concentrations were determined using a validated LC-MS/MS method. Pharmacokinetic parameters were calculated by using non-compartmental analysis. Comparisons of levocetirizine pharmacokinetics were conducted with maximum concentration (C max) and the area under the plasma concentration-time curve from dosing until 48 h post-dose (AUC0-48) after each treatment.Clinical trial registration numberClinicalTrials.gov identifier is NCT01622283.ResultsThe mean C max and AUC0-48 of levocetirizine after a single dose of levocetirizine OS 5 mg and cetirizine DS 10 mg were 203.3 ± 42.49 ng/mL and 1814.9 ± 304.22 ng.hr/mL, and 196.5 ± 31.31 ng/mL and 1710.5 ± 263.31 ng hr/mL, respectively. The ratios and the 90% CIs of the geometric least squares means ratios of C max and AUC0-48 were 1.027 (0.968-1.091) and 1.059 (1.024-1.094), respectively.LimitationThe small sample size and single dose design of this study prevent definitive conclusions regarding the pharmacokinetics and safety of levocetirizine OS in a Japanese patient population being made. Study limitations include conducting the study in adult males, not in children.ConclusionsLevocetirizine exposure in plasma was equivalent when given as levocetirizine OS 5 mg and as cetirizine DS 10 mg. Both preparations were safe and well-tolerated in healthy Japanese male subjects.

Project description:A large problem of histamine receptor research is data heterogeneity. Various experimental approaches, the complex signaling pathways of mammalian cells, and the use of different species orthologues render it difficult to compare and interpret the published results. Thus, the four human histamine receptor subtypes were analyzed side-by-side in the Sf9 insect cell expression system, using radioligand binding assays as well as functional readouts proximal to the receptor activation event (steady-state GTPase assays and [35S]GTP?S assays). The human H1R was co-expressed with the regulators of G protein signaling RGS4 or GAIP, which unmasked a productive interaction between hH1R and insect cell G?q. By contrast, functional expression of the hH2R required the generation of an hH2R-Gs? fusion protein to ensure close proximity of G protein and receptor. Fusion of hH2R to the long (Gs?L) or short (Gs?S) splice variant of G?s resulted in comparable constitutive hH2R activity, although both G protein variants show different GDP affinities. Medicinal chemistry studies revealed profound species differences between hH1R/hH2R and their guinea pig orthologues gpH1R/gpH2R. The causes for these differences were analyzed by molecular modeling in combination with mutational studies. Co-expression of the hH3R with G?i1, G?i2, G?i3, and G?i/o in Sf9 cells revealed high constitutive activity and comparable interaction efficiency with all G protein isoforms. A comparison of various cations (Li+, Na+, K+) and anions (Cl-, Br-, I-) revealed that anions with large radii most efficiently stabilize the inactive hH3R state. Potential sodium binding sites in the hH3R protein were analyzed by expressing specific hH3R mutants in Sf9 cells. In contrast to the hH3R, the hH4R preferentially couples to co-expressed G?i2 in Sf9 cells. Its high constitutive activity is resistant to NaCl or GTP?S. The hH4R shows structural instability and adopts a G protein-independent high-affinity state. A detailed characterization of affinity and activity of a series of hH4R antagonists/inverse agonists allowed first conclusions about structure/activity relationships for inverse agonists at hH4R. In summary, the Sf9 cell system permitted a successful side-by-side comparison of all four human histamine receptor subtypes. This chapter summarizes the results of pharmacological as well as medicinal chemistry/molecular modeling approaches and demonstrates that these data are not only important for a deeper understanding of HxR pharmacology, but also have significant implications for the molecular pharmacology of GPCRs in general.

Dataset Information

Differential Regulation of Thermodynamic Binding Forces of Levocetirizine and (S)-Cetirizine by Lys191 in Human Histamine H₁ Receptors.

Publications

Differential Regulation of Thermodynamic Binding Forces of Levocetirizine and (<i>S</i>)-Cetirizine by Lys191 in Human Histamine H₁ Receptors.

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