Histamine H1-receptor

Eosinophils are well known to play essential roles in the development and maintenance of allergic diseases. However, the influence of histamine H1 receptor antagonists on eosinophil functions, especially chemokine production, are not well-defined. Therefore, in the present study, we examined the influence of histamine H1 receptor antagonist on chemokine production by eosinophils through the use of levocetirizine in vitro and in vivo. Eosinophils prepared from mice were stimulated with specific antigens in the presence of different concentrations of levocetirizine. After 24 h, regulated on activation normal T cell expressed and secreted (RANTES) and eotaxin levels in culture supernatants were measured by enzyme-linked immunosorbent assay (ELISA). Patients with Japanese cedar pollinosis were treated with 5 mg levocetirizine once a day for four weeks during the pollen season (February 2012 to April 2012). RANTES and eotaxin levels in nasal secretions were also examined by ELISA. The addition of levocetirizine to eosinophil cultures caused a dose-dependent decrease in the ability of cells to produce RANTES and eotaxin in response to antigen stimulation, and the minimum concentration that caused a significant decrease was 0.05 μM. Although cetirizine also exerted suppressive effects on the production of RANTES and eotaxin by eosinophils, the minimum concentration that caused significant suppression was 0.15 μM, which was three-times higher than that of levocetirizine. Oral administration of levocetirizine for four weeks also reduced RANTES and eotaxin levels in nasal secretions from patients with pollinosis, along with attenuation of clinical symptoms. The ability of levocetirizine to reduce RANTES and eotaxin levels may account, at least in part, for the clinical efficacy of the agent for allergic disorders, including allergic rhinitis[2].

Male Sprague-Dawley rats treated with levocetirizine (0.4 mg/kg; oral) had Cmax, AUC0-t, AUC0-∞, and t1/2 values of 0.34 μg/mL, 3.26 μg h/mL, and 3.67 μg, correspondingly. 2.34 hours and h/mL in Sprague-Dawley rats, respectively [1].

Assay for transcription factor activation. NF-κB activity in cultured eosinophils was analyzed by commercially available NF-κB ELISA test kits that contained sufficient reagents and a monoclonal antibody against p65, according to the manufacturer's recommendations. In brief, nuclear extract (5.0 mg of protein) from eosinophils was introduced into each well of 96-well microplates precoated with oligonucleotide containing NF-κB consensus site (5’-GGGACTTTCC-3’) in a volume of 20.0 μl, followed by incubation for one hour at 25°C. After washing three times, 100 μl of monoclonal antibody against p65 was added to the appropriate wells and incubated for a further one hour at 25°C. Anti-IgE horseradish peroxidase (HRP) conjugate in a volume of 100 μl was then added and the plates incubated for a further one hour at 25°C. The absorbance at 450 nm was measured after the addition of tetramethylbenzidine (TMB) solution. AP-1 activity was also measured with commercially available AP-1 ELISA test kit in a similar manner[2].

Assay for mRNA expression. Poly A+ mRNA was separated from cultured cells with oligo(dT)-coated magnetic micro beads. The first-strand cDNA was synthesized from 1.0 mg of PolyA+ mRNA using a Superscript cDNA synthesis kit according to the manufacturer's instructions. Polymerase chain reaction (PCR) was then carried out using a GeneAmp 5700 Sequence Detection Syste. The PCR mixture consisted of 2.0 μl of sample cDNA solution (100 ng/μl), 25.0 μl of SYBR-Green Mastermix, 0.3 μl of both sense and antisense primers, and distilled water to give a final volume of 50.0 μl. The reaction was conducted as follows: 4 min at 94°C, followed by 40 cycles of 15 s at 95°C and 60 s at 60°C. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was amplified as an internal control. mRNA levels were calculated by using the comparative parameter threshold cycle and normalized for GAPDH. The nucleotide sequences of the primers were as follows: for RANTES, 5’-CCTCACCATCATCCTCACTGCA-3’ (sense) and 5’-TCTTCTCTGGGTTGGCACACAC-3’ (antisense), for eotaxin, 5’-CCCTTTTCTGTTCTGCTGACAAG-3’ (sense) and 5’-GAAGAGTCCCTCGATGTGGCTA-3’ (antisense), and for GAPDH, 5’-GTCTTCTGGGTGGCAGTGAT-3’ (sense) and 5’-CCCTTT TCTGTTCTGCTGACAAG-3’ (antisense)[2].

Animal/Disease Models: 30 male SD (SD (Sprague-Dawley)) rats (8 weeks old; 200-250 g) [1]
Doses: 0.4 mg/kg
Route of Administration: Oral (pharmacokinetic/PK/PK analysis)
Experimental Results: Cmax, AUC0-t, AUC0-∞ and t1 /2 were 0.34μg/mL, 3.26μg·h/mL, 3.67μg·h/mL and 2.34 hrs (hrs (hours)) respectively.

[1]. Lohar P, et al. Simultaneous bioanalysis and pharmacokinetic interaction study of acebrophylline, levocetirizine and pranlukast in Sprague-Dawley rats. Biomed Chromatogr. 2019 Dec;33(12):e4672.
[2]. Levocetirizine Pretreatment Mitigates Lipopolysaccharide-Induced Lung Inflammation in Rats. Biomed Res Int. 2018 Aug 13;2018:7019759.

Levocetirizine Dihydrochloride is the dihydrochloride salt form of the active levorotatory enantiomer of cetirizine, levocetirizine; a third generation, non-sedating, selective histamine H1 receptor antagonist, with antihistamine, anti-inflammatory and potential anti-angiogenic activities. Levocetirizine competes with endogenous histamine for binding at peripheral H1-receptor sites on the effector cell surface. This prevents the negative symptoms associated with histamine release and an allergic reaction. In addition, as histamine plays an important role in angiogenesis during an allergic inflammatory reaction, blocking the action of histamine may modulate the expression of proangiogenic factors and thus may prevent angiogenesis. As a third-generation histamine H1 receptor antagonist, levocetirizine has fewer side effects than most second-generation antihistamines.

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The combination of acebrophylline (ABP), levocetirizine (LCZ) and pranlukast (PRN) is used to treat allergic rhinitis, asthma, hay-fever and other conditions where patients experience difficulty in breathing. This study was carried out with the aim of developing and validating a reverse-phase high-performance liquid chromatographic bioanalytical method to simultaneously quantitate ABP, LCZ and PRN in rat plasma. The objective also includes determination of the pharmacokinetic interaction of these three drugs after administration via the oral route after individual and combination treatment in rat. Optimum resolution between the analytes was observed with a C18 Kinetex column (250 mm × 4.6 mm × 5 μm). The chromatography was performed in a gradient elution mode with a 1 mL/min flow rate. The calibration curves were linear over the concentration range of 100-1600 ng/mL. The intra- and inter-day precision and accuracy were found to be within acceptable limits as specified in US Food and Drug Administration guideline for bioanalytical method validation. The analytes were stable on the bench-top (8 h), after three freeze-thaw cycles, in the autosampler (8 h) and as a dry extract (-80°C for 48 h). The statistical results of the pharmacokinetic study in Sprague-Dawley rats showed a significant change in pharmacokinetic parameters for PRN upon co-administration of the three drugs.[1]

C21H27CL3N2O3

461.8

460.1087

C, 54.62; H, 5.89; Cl, 23.03; N, 6.07; O, 10.39

130018-87-0

Cetirizine;83881-51-0;Cetirizine dihydrochloride;83881-52-1;Cetirizine-d4;1219803-84-5;Cetirizine-d8;774596-22-4;Levocetirizine;130018-77-8;Cetirizine-d4 dihydrochloride;Cetirizine-d8 dihydrochloride;2070015-04-0

9955977

Typically exists as white to off-white solids at room temperature

542.1ºC at 760 mmHg

215-220ºC

281.6ºC

4.63

53.01

C1=CC=C(C=C1)[C@H](C2=CC=C(C=C2)Cl)N3CCN(CC3)CCOCC(=O)O.Cl.Cl

PGLIUCLTXOYQMV-GHVWMZMZSA-N

InChI=1S/C21H25ClN2O3.2ClH/c22-19-8-6-18(7-9-19)21(17-4-2-1-3-5-17)24-12-10-23(11-13-24)14-15-27-16-20(25)26;;/h1-9,21H,10-16H2,(H,25,26);2*1H/t21-;;/m1../s1

2-[2-[4-[(R)-(4-chlorophenyl)-phenylmethyl]piperazin-1-yl]ethoxy]acetic acid;dihydrochloride

UCB28556; UCB 28556; Levocetirizine dihydrochloride; 130018-87-0; Xyzal; (R)-Cetirizine dihydrochloride; Levocetirizine HCl; Xusal; UNII-SOD6A38AGA; SOD6A38AGA; (-)-Cetirizine;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~100 mg/mL (~216.54 mM)
H2O : ~100 mg/mL (~216.54 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.41 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (5.41 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (5.41 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 50 mg/mL (108.27 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.

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 (Please use freshly prepared in vivo formulations for optimal results.)