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The interaction between antihistaminic drug oxatomide (OXT) and calf-thymus DNA (CT-DNA) has been investigated in a physiological buffer (pH 7.4) using UV–Vis, fluorescence, 1H NMR and circular dichroism spectral techniques coupled with viscosity measurements, KI quenching, voltammetry and in silico molecular modeling studies. OXT binds with CT-DNA in a concentration-dependent manner. At a lower [Drug]/[CT-DNA] molar ratio (0.6–0.1), OXT intercalates into the base pairs of CT-DNA, while at a higher [Drug]/[CT-DNA] molar ratio (13–6), the drug binds in the minor grooves of CT-DNA. The binding constants for the interaction are found to be in the order of 103–105 M−1, and the groove binding mode of interaction exhibits a slightly higher binding constant than that of intercalative mode. Thermodynamic analysis of binding constants at three different temperatures suggests that both these modes of binding are mainly driven by hydrophobic interactions (ΔHo > 0 and ΔSo > 0). Voltammetric investigations indicate that the electro-reduction of OXT is an adsorption controlled process and shifts in reduction peak potentials reiterate the concentration-dependent mode of binding of the drug with CT-DNA. The free energy landscape obtained at the all-atom level, using metadynamics simulation studies, revealed two major binding forces: partial intercalation and minor groove binding, which corroborate well with the experimental results. Communicated by Ramaswamy H. Sarma

Histamine H1 receptor (H1R) expression influences the severity of allergy symptoms. We examined the effect of inverse agonists on H1R gene expression. Two inverse agonists (carebastine and mepyramine), but not the neutral antagonist oxatomide, decreased inositol phosphate accumulation. The inverse agonists also decreased H1R gene expression and down-regulated H1R mRNA below basal expression, while basal H1R mRNA expression was maintained after oxatomide treatment. These results suggest that inverse agonists more potently alleviate allergy symptoms by not only inhibiting stimulus-induced up-regulation of H1R gene expression but also by suppressing basal histamine signaling through their inverse agonistic activity. Keywords:: constitutive activity, histamine H1 receptor gene transcription, inverse agonist

A rapid, sensitive and specific high-performance liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) method has been developed and validated for the determination of oxatomide in human plasma. Flunarizine hydrochloride was employed as the internal standard (IS). The analytes were chromatographically separated on a Shimadzu Shim-pack VP-ODS C18 column (250 x 2.0 mm i.d.) with a mobile phase consisting of methanol and aqueous ammonium acetate solution (10 mm, pH 4.0; 85:15, v/v). Detection was performed on a single quadrupole mass spectrometer using an electrospray ionization interface with the selected-ion monitoring (SIM) mode. The method showed excellent linearity (r = 0.9995) over the concentration range of 0.5-500 ng/mL with good accuracy and precision. The intra- and inter-batch precisions were within 10% relative standard deviation. The recoveries were more than 90%. The validated method was successfully applied to a preliminary pharmacokinetic study of oxatomide in Chinese healthy male volunteers.

Objective: Inflammatory mediators (IMs) play a major role in the production of middle ear effusion (MEE). Tumor necrosis factor (TNF)-α and leukotrienes (LTs) appear to be important in the pathogenesis of otitis media with effusion (OME). The purpose of this study is to determine the effect of TNF-α and LT antagonist on the outcome of experimental immune-mediated OME. Study Design: Prospective. Methods: Otitis media was induced in rats by injecting keyhole limpet hemocyanin (KLH) transtympanically 7 days after systemic immunization. Experimental groups were treated with soluble TNF receptor type I (sTNF RI) or oxatomide simultaneously. Seventy-two hours after transtympanic injection, MEE was aspirated, and temporal bone was taken. Vascular permeability (VP) of the middle ear mucosa was measured using the Evans blue dye technique. Hematoxylin-eosin stain and immunohistochemical stain for leukocyte common antigen was performed. Results: In KLH, sTNF RI, and oxatomide groups, MEE was developed in 83%, 0%, and 66% of the ears, respectively. The sTNF RI group showed significant decrease in effusion production, inflammation, mucosal thickening, and VP compared with the KLH group. These parameters were less significant in the oxatomide group than in the sTNF RI group. Conclusion: Transtympanic administration of sTNF RI and oxatomide appears to suppress the development of immune-mediated MEE.

The effects of anti-pruritic drugs on scratching behavior associated with passive cutaneous anaphylaxis in histamine H(1) receptor-deficient and wild-type mice were studied. Passive sensitization with mouse monoclonal anti-dinitrophenyl-immunoglobulin E (IgE) resulted in an increase in the incidence of scratching behavior induced by intravenous injection of dinitrophenyl-ovalbumin in both wild-type and histamine H(1) receptor-deficient mice. The histamine H(1) receptor antagonist diphenhydramine inhibited scratching behavior induced by antigen in passively sensitized wild-type mice, whereas no effect was observed in histamine H(1) receptor-deficient mice. On the other hand, oxatomide inhibited scratching behavior in both mice, although the effect in wild-type mice was more potent than that in histamine H(1) receptor-deficient mice. Tranilast inhibited scratching behavior with the same potency in both mice. We concluded that the scratching behavior associated with passive cutaneous anaphylaxis involves not only histamine H(1) receptors but also other chemical mediators. Furthermore, the results of the present study indicated that oxatomide has an antagonistic effect on histamine H(1) receptors as well as anti-pruritic effect in vivo.

A rapid liquid chromatographic method was developed for the determination of oxatomide in its finished active pharmaceutical ingredient form and in the presence of its process impurities. The method was developed on a sub 2 mm Hypersil Zorbax XDB C18 column (30 3 4.6 mm, i.d., 1.8 mm). The rapid method employed a gradient mobile phase consisting of solvent A: 0.01 M tetrabutylammonium hydrogen sulfate and 0.5% (w/v) ammonium acetate in water and solvent B: acetonitrile. A flow rate of 2 mL/min was employed with the diode-array detector set at 230 nm. The original method supplied by Janssen Pharmaceuticals Ltd was run on a Thermo Scientific octadecylsilyl silica gel C18 column (100 3 4.6 mm, i.d., 5 mm) with an analysis time of 20 min. The main aim was to substantially reduce the analysis time while maintaining good efficiency. Run-time was reduced to 6.5 min with a total loss in analysis time of 68%. Solvent consumption was also reduced by 68%. Validation according to the International Conference of Harmonization guidelines was undertaken. The parameters examined were accuracy, precision, linearity, selectivity, robustness, limit of detection and limit of quantification; all criteria were met. Sample stability testing was also carried out. Oxatomide proved stable under ambient and 48C temperatures and in the presence of light for up to 24 h.

Experimental and epidemiological studies have indicated that central histaminergic neuron system plays an important role in inhibition of convulsive disorders through histamine H(1)-receptors, especially in developing period. Histamine H(1) antagonists increase the duration of electrically induced convulsions in 21-day-old mice, but not in 42-day-old mice. Epidemiological studies suggested that histamine H(1) antagonist may be one of the risk factors in febrile convulsions. In histidinemic patients who were considered to have high brain histamine content, the incidence of convulsions was lower than that of ordinary population.The centrally acting histamine H(1) antagonists including pyrilamine and ketotifen facilitate the development of amygdaloid kindling in rats, an experimental model of epileptogenic process. On the contrary, epinastine, a histamine H(1) antagonist which scarcely enters the brain, shows no facilitation. These findings suggest that the central histaminergic neuron system plays an inhibitory role on the seizure development through central histamine H(1)-receptors.Recently, three cases has been reported in which West syndrome developed 8-10 days after ketotifen or oxatomide administration. Considering experimental and clinical studies, histamine H(1) antagonists may be associated with West syndrome and may be hazardous to infants. Further careful experimental and clinical studies will be required to elucidate the relationships between West syndrome and central histaminergic neuron system.

Background: Lipid mediators play a pivotal role in the pathogenesis of allergic and inflammatory reactions. These molecules include metabolites of arachidonic acid (AA) and the group of platelet-activating factor (PAF)-related phospholipids. The initial step in the synthetic pathway of both classes of lipid mediators is catalyzed by members of the phospholipase A2 (PLA2) family. Oxatomide is a histamine H1 receptor antagonist currently used in the treatment of allergic disorders. Preliminary evidence indicates that oxatomide may exert anti-inflammatory activities unrelated to H1 receptor antagonism. Methods: We investigated the effect of oxatomide on lipid mediator production by human neutrophils. Results: Preincubation (15 min, 37°C) of neutrophils with oxatomide (10–100 µM) concentration-dependently inhibited (10–70%) the release of AA induced by the Ca2+ ionophore A23187 (0.5 µM). Oxatomide also comparably inhibited the release of the four major metabolites of AA induced by A23187 (LTB4, 20-OH-LTB4, 20-COOH-LTB4 and 5-HETE). In addition, oxatomide reduced by 60% the production of PAF induced by A23187. The simultaneous inhibition of the production of AA metabolites and PAF suggested that oxatomide could influence the activity of cytosolic PLA2 (cPLA2). To test this hypothesis, we evaluated the functional activity of cPLA2 in neutrophils preincubated with oxatomide. This preincubation inhibited (72 ± 24%) the increase in cPLA2 activity induced by A23187. Conclusions: These results indicate that oxatomide reduces the biosynthesis of lipid mediators in human neutrophils by inhibiting cPLA2. This inhibitory effect of oxatomide may contribute to the anti-inflammatory activity of this drug in allergic diseases.

Anti-scratch effects of oxatomide and epinastine were examined in mice. Scratching behavior and cutaneous reactions were induced in BALB/c, ICR and ddY mice by dinitrofluorobenzene painting, passive cutaneous anaphylaxis and substance P injection, respectively. Although oxatomide and epinastine failed to inhibit scratching behavior in BALB/c mice, they inhibited the cutaneous reaction significantly. The drugs potently inhibited both scratching behavior and cutaneous reaction in ICR mice. They also inhibited scratching behavior and cutaneous reaction in ddY mice, although cetirizine and terfenadine failed to affect them. Histamine did not induce frequent scratching behavior in BALB/c and ddY mice. These results indicate that oxatomide and epinastine inhibit the scratching behavior in ICR mice associated with passive cutaneous anaphylaxis mainly through an antagonistic action on histamine H 1 receptors. The results also indicate that these drugs inhibit substance P-induced scratching behavior in ddY mice through an action independent of the antagonistic action on histamine H 1 receptors.

Our previous study revealed that passive cutaneous anaphylaxis (PCA) can be produced in congenitally mast cell-deficient WBB6F1-W/Wv (abbreviated as W/Wv) mice on sensitization with undiluted or slightly diluted allogeneic and xenogeneic antisera but not on sensitization with allogeneic monoclonal immunoglobulin (Ig)E and IgG1 antibodies regardless of the antibody concentration [1]. In view of these findings, the present study was conducted to characterize PCA in this strain from its drug susceptibilities using mast cell-bearing WBB6F1-(+)/+ (abbreviated as +/+) and B6D2F1 mice as references. PCA in W/Wv mice mediated by a low dilution (1:4) of hyperimmune serum to bovine serum albumin of the B6D2F1 mouse origin was markedly suppressed by CV-6209, an antagonist of platelet-activating factor (PAF), but not by antihistamines such as cyproheptadine and oxatomide. In contrast, PCA in +/+ and B6D2F1 mice mediated by a high dilution (1:128) of the anti-serum (virtually by IgG1 antibody) was nearly completely suppressed by antihistamines but not by CV-6209. A remarkable difference between PCA in W/Wv and reference mice was also observed in the susceptibility to monoclonal anti-mouse granulocyte (Gr-1) antibody: PCA in W/Wv mice was potently suppressed by the 1- to 3-day pretreatment with this antibody but that in references was not at all. Putting these present results together with the previous finding that anti-granulocyte antibody greatly reduces circulatory Gr-1+ leukocytes, 1 to 3 days after the treatment [2], it is highly probable that PCA in W/Wv mice mediated by some antibody isotypes other than IgE and IgG1 is produced by PAF mainly released from Gr-1+ cells, while IgG1 antibody-mediated PCA in mast cell-bearing reference mice is evoked by histamine derived from mast cells. PCA homologous to that in W/Wv mice could also be produced in the reference mice on sensitization with undiluted or slightly diluted antiserum, when generalized blueing due to excess IgG1 antibody was removed by the oxatomide treatment before the antigen challenge.

While it is clear that the clinical expression of IgE-mediated diseases depends upon the actions of multiple mediators, histamine, the earliest recognized mediator of allergy, remains a prominent contributor. Histamine released from mast cells binds to specific receptors (H1, H2, H3) to produce its clinical effects. The cardinal features of asthma include smooth muscle spasm, mucosal edema, inflammation and mucus secretion. It has been demonstrated that two of these features, bronchospasm and mucosal edema, can be caused by H1-receptor stimulation, while H2- and possibly H1-activation are probably minor causes of mucus secretion. Histamine interacts directly with the endothelial cells (EC) and induces permeability, a transient expression of P-selectin and the secretion of lipid mediators (e.g. PGI2, PAF and LTB4). Moreover, histamine induces a significant increase of IL-6 and IL-8 secretion by EC. Since IL-8 exerts a chemotactic activity for neutrophils, eosinophils and basophils, and IL-6 is involved in endothelium permeability, the secretion of cytokines may be involved in the late phase reaction. Some antihistamines (i.e., levocabastine, terfenadine, loratadine, azelastine and oxatomide) can reduce ICAM-1 expression. The participation of histamine in the allergic inflammation, including asthma, must be re-examined, since the effects of histamine are more widespread.

The effects of a newly synthesized compound, SWR-00151 (4-[2-[4-(1H-indol-3-yl)-1-piperidinyl)ethyl]-2(1H)-quinolinone), on experimental Type I allergic models were investigated. Results obtained were as follows: The compound (3 approximately 30 mg/kg, p.o.) dose-dependently inhibited 48-hr passive cutaneous anaphylaxis (PCA) in the rat. From the strong antagonism against the histamine-induced contraction of the isolated guinea pig ileum and the lack of suppressive effect on anaphylactic histamine release from rat peritoneal exudate cells, it is deduced that the compound's inhibitory action against PCA is due to antihistaminic action. Both gamma 1-rich serum- and IgE-rich serum-mediated experimental asthmas in the guinea pig were also considerably inhibited by a small dose (1 mg/kg, p.o.) of the compound. The inhibitory mechanism seems to be almost the same as that of the PCA because the compound did not show any effect on the experimental asthma in guinea pigs pretreated with H1- and H2-antihistaminics. In addition to that, it is well known that the model is largely mediated by anaphylactically released histamine. On the other hand, while ketotifen and oxatomide, which possess potent antihistaminic activity, modestly suppressed a rat experimental asthma SWR-00151 still demonstrated a substantial inhibitory activity, strongly suggesting that histamine does not play an important role in this asthma model. Serotonin was revealed to be partly responsible for the early phase of the reaction by the assessment with methysergide, an antiserotonergic, and SWR-00151 as well as oxatomide and ketotifen showed slight antagonism against serotonin in high concentrations (10(-6) and 10(-5) M) in vitro. When thromboxane (TX) B2 in the plasma was measured during the reaction, significant increased levels of the chemical mediator were found, which were obviously prevented by the treatment with SRW-00151. From these results, SWR-00151 is expected to be a drug effective for the treatment of asthma through mechanisms not only of antihistaminic action but also through inhibition of anaphylactic formation/release of other mediators like TXA2.

Summary Background TMK688 is being developed as ati anti-allergic drug having both 5-lipoxygenasc inhibitory activity atid anti-histaminc activity. Method We compared the inhibition ol the late asthmatic rcsponses by TMK688 with that by other anti-allergic agents in actively sensitized guinea pigs, and examined the relationship between 5-lipoxygetiase inhibition and the late asthmatic responses Results At 1–3.2 mg/kg, TMK688 inhibited the increases in respiratory resistance, leukotriene (LT) B4 and C4 production in the lungs and eosinophil infiltralion into the alveoli during the late asthmatic response, whereas the eliects tended to lessen at the dose of 10mg/kg. These effects are thought to be caused by the 5-lipoxygenase inhibitory activity of TMK688 because Azelastine. an anti-allergic drug having potent antihistamine activity, exhibited no effect. ONO-1078. a peptide LT antagonist. inhibited the late-phase bronchoconstriction at a dose of 100mg/kg p.o., but not the increase in the infiltration of inflammatory cells into the alveoli, suggesting that the late-phase bronchoconstriction is induced, in part, by peptide LTs, i.e. LT C4, D4 and E4 atid that the inflammatory cell infiltration may be caused by LTB4, TMK688 inhibited the immediate bronchoconstriction dose-dependently, and the effect was significant at a dose of 10mg/kg orally. Since Azelastine, Ketotifen and Oxatomide suppressed the bronchoconstriction at far lower doses than did TMK688, the inhibitory effect was mainly caused by its antihistamine activity. Conclusions TMK688 appears to be a novel anti-allergic drug having inhibitory effects on both the bronchoconstriction and the infiltration of inflatnmutory cells during late asthmatic responses.

Oxatomide (OXA), a histamine H1 receptor antagonist, is effective in the treatment of patients with allergic rhinitis, some allergic skin disorders, and bronchial asthma. We have characterized the effect of OXA on the immunologic release of preformed (histamine and tryptase) and de novo synthesized mediators (leukotriene C4:LTC4 and prostaglandin D2:PGD2) from human basophils and mast cells purified (from 10 to 82%) from human lung parenchyma (HLMC) and skin tissue (HSMC). Preincubation (15 min, 37 degrees C) of basophils with OXA (10(-7)-10(-5) M) before Der p I antigen or anti-IgE challenge concentration-dependently (10-40%) inhibited the immunologic release of histamine and LTC4. OXA (10(-7)-10(-5) M) also inhibited (10-40%) histamine, tryptase and LTC4 release from HLMC activated by anti-IgE. In addition, OXA caused a concentration-dependent inhibition of histamine, tryptase and PGD2 release from HSMC immunologically challenged with a monoclonal antibody against the alpha chain of the high affinity receptor for IgE (anti-Fc epsilon RI) or anti-IgE. These results demonstrate that OXA exerts anti-inflammatory activities by inhibiting the release of preformed and de novo synthesized mediators from human basophils and mast cells.

We studied the effects of antiallergic drugs, epinastine, ketotifen, oxatomide, mequitazine and cromolyn sodium on superoxide anion (O 2 - ) generation from rat neutrophils. Epinastine, ketotifen, oxatomide and mequitazine dose-dependently prevented the N-formyl-Met-Leu-Phe- and phorbol 12-myristate 13-acetate-induced O 2 - generation, but cromolyn sodium did not prevent it. When membrane and cytosol fractions were incubated with each drug, epinastine, ketotifen and mequitazine prevented O 2 - generation. On the other hand, when only the membrane fraction was incubated with each drug, ketotifen and mequitazine prevented O 2 - generation, but epinastine did not. Epinastine may inhibit the NADPH oxidase system through the obstruction of NADPH oxidase-associated cytosol components.

According to the previous concepts of antiallergic drugs, it was difficult to select the essential antiallergic drugs which was necessary for treatment of patients with allergy. We, therefore, attempted to discover that antiallergic drugs have the any pharmacologic actions on the process of allergic response. We divided into three stages on the process of allergic response in this animal experiment. Antiallergic drugs used in this study were azelastine hydrochloride, ketotifen fumarate, tranilast and oxatomide. First, we observed the suppression of IgE production in rats on the administration of azelastine hydrochloride. Secondly, we observed that azelastine hydrochloride and ketotifen fumarate inhibited the interaction between anti-DNP-As and DNP-As on ELISA. Thirdly, we observed that tranilast and azelastine hydrochloride reduced, more effectively than any other antiallergic drug, the release of histamine from rabbit washed platelets by PAF.In addition, it was observed that azelastine hydrochloride and ketotifen inhibited dose-dependently the increase of permeability induced by PAF or histamine in rats. From these results, we could propose the new concepts that some of antiallergic drug inhibited the IgE production.

1. Guinea-pig tracheal contractions by histamine and by the cholinoceptor agonist, arecoline, are significantly enhanced (30% and 20%, respectively), 96 h after intra-tracheal inoculation with Parainfluenza-3 (PI-3) virus. 2. The airway hyperresponsiveness in animals inoculated with virus coincides with a significant increase in the number of broncho-alveolar cells (82%), and in the albumin concentration (121%) in lung lavage fluid, relative to values obtained in guinea-pigs challenged with control solution. 3. The chemiluminescence production by isolated broncho-alveolar cells, obtained from virus-infected guinea-pigs 96 h after inoculation stimulated with PI-3 virus in vitro, is significantly reduced by 42% relative to broncho-alveolar cells obtained from animals inoculated with control solution. This diminution was not specific for stimulation by PI-3 virus since the chemiluminescence production was also significantly reduced by 30% in response to zymosan. 4. Pretreatment of the guinea-pigs with the anti-allergic drugs, oxatomide (2.5 mg kg-1) or nedocromil (2.5 mg kg-1), or the specific H1-histamine receptor antagonist, levocabastine (0.25 mg kg-1), administered intra-peritoneally twice a day for five successive days, inhibits the virus-induced airway hyperresponsiveness, suppresses the influx of broncho-alveolar cells and increase in albumin content, and corrects the reduced chemiluminescence production by broncho-alveolar cells in response to zymosan. 5. In contrast, the cyclo-oxygenase inhibitor, suprofen (5.0 mg kg-1), the 5-HT2 receptor antagonist, ketanserin (0.63 mg kg-1), or the Ca2+ overload blocker, flunarizine (2.5 mg kg-1) do not modify the above mentioned processes. 6. The platelet-activating factor receptor antagonist, WEB 2170 (10 mg kg-1), reduces virus-induced airway hyperresponsiveness and influx of broncho-alveolar cells into the lungs but does not attenuate the increase of albumin in the bronchial lavage fluid. 7. Guinea-pigs nebulized with histamine, twice a day (30 min) during 4 successive days, do not demonstrate an increased airway responsiveness, but instead show tachyphylaxis in response to histamine in vitro. In addition, no influx of inflammatory cells is found in these animals. 8. These results suggest that histamine does not directly increase the responsiveness of the guinea-pig trachea; however, histamine may be involved in a cascade of events leading to airway hyperresponsiveness after a viral infection, a process that could be related to an influx and/or an activation of broncho-alveolar cells after PI-3 virus stimulation.

A number of N-[4-[4-(1H-indol-3-yl)piperidinoalkyl]-2- thiazolyl]alkanesulfonamides (8--21) were synthesized and evaluated for their preventive effects on systemic anaphylaxis in guinea pigs. Structure-activity analysis revealed that methane- and ethanesulfonamide derivatives having a one to three methylene tether between the piperidine and thiazole rings exhibited potent activity but the introduction of a substituent on the indole part reduced the activity. Administration (100 mg/kg p.o.) of the four compounds 8, 9, 12, 13, together with ketotifen, oxatomide, terfenadine and azelastine as reference compounds, to mice revealed that only compound 8 caused no significant increase of the sleeping time induced by hexobarbital. In addition, compound 8 (10 mg/kg i.v.) did not change the electroencephalogram in conscious rabbits. These results led to the selection of N-[4-[4-(1H-indol-3-yl)piperidinomethyl]-2-thiazolyl]methanesulfon amide (8, FK613) for further development as a novel antiallergic agent. Clinical evaluation of FK613 is now in progress.

The molecular complexes of the donors ketaconazole (KTZ) and oxatomide (OXA) drugs with 2,3,5,6-tetrachloro-1,4-benzoquinone (p-chloranil, p-CHL) have been investigated spectroscopically (UV–vis, FT-IR and 1H NMR) and spectrofluorimetrically in different solvents and temperatures. The stoichiometry of the complexes was found to be 1:1. The data are discussed in terms of formation constant, molar extinction coefficient, oscillator strength, dipole moment, ionization potential, dissociation energy and thermodynamic parameters. The results indicated that the formation of molecular complex is spontaneous and endothermic. The fluorescence quenching studies indicated that the interaction of the donors is spontaneous and the fluorescence quenching increased with an increase in the intensity of complexation with the acceptor.

A number of N-[4-[4-(1H-indol-3-yl)piperidinoalkyl]-2-thiazolyl]alkanesulfonamides (8-21) were synthesized and evaluated for their preventive effects on systemic anaphylaxis in guinea pigs. Structure-activity analysis revealed that methane- and ethanesulfonamide derivatives having a one to three methylene tether between the piperidine and thiazole rings exhibited potent activity but the introduction of a substituent on the indole part reduced the activity. Administration (100mg/kg p.o.) of the four compounds 8, 9, 12, 13, together with ketotifen, oxatomide, terfenadine and azelastine as reference compounds, to mice revealed that only compound 8 caused no significant increase of the sleeping time induced by hexobarbital. In addition, compound 8 (10mg/kg i.v.) did not change the electroencephalogram in conscious rabbits. These results led to the selection of N-[4-[4-(1H-indol-3-yl)piperidinomethyl]-2-thiazolyl]methanesulfonamide (8, FK613) for further development as a novel antiallergic agent. Clinical evaluation of FK613 is now in progress.

At 1-week intervals, 12 healthy volunteers received oral cetirizine 10mg, oxatomide 30mg, ketotifen 1mg or placebo under randomised double-blind crossover conditions. Wheal and flare were induced by skin prick tests using histamine 100 and 500 g/L before, and 4, 8 and 24 hours after, drug ingestion. At each session volunteers recorded levels of pruritus and sedation on two 10cm visual analogue scales. Cetirizine was significantly more potent than oxatomide (p = 0.0001), ketotifen (p = 0.0002) or placebo (p = 0.0001) in the inhibition of wheals, with similar results for flare (p = 0.0007, 0.0001 and 0.0002, respectively). Relief from pruritus was significantly better with cetirizine than with oxatomide (p = 0.003) or ketotifen (p = 0.002). Cetirizine was slightly better than placebo in relieving pruritus, but this was not statistically significant. Oxatomide and ketotifen were clearly more sedative than cetirizine (p = 0.05 and p = 0.0006, respectively), which was not significantly different from placebo. In conclusion, cetirizine appeared to be better in reducing histamine-induced wheal, flare and pruritus compared with ketotifen and oxatomide, and had less sedative effect.

Inhibitory effects of various histamine receptor-blocking agents including antiallergic agents on metabolic activations of neutrophils were examined by measuring leukotrienes (C4, D4 and E4) formation, arachidonic acid release and superoxide generation. The stimulation of neutrophils by calcium ionophore (A23187) causes the production of leukotrienes concomitantly with the release of arachidonic acid from the cells and these were effectively diminished with a variety of antihistaminic agents. Almost all the histamine H1 receptor-blocking drugs studied here showed as inhibitors of the metabolic activations of neutrophils, although the degree was dependent on the drug concentrations. The order of potency of the inhibitory effects on the arachidonic acid release were: homochlorcyclizine, clemastine, and azelastine (IC50 less than 20 microM) greater than oxatomide (IC50 less than 60 microM) and diphenylpyraline greater than triprolidine, meclizine, diphenhydramine (IC50 greater than 100 microM). The superoxide generation from neutrophils activated by phorbol 12-myristate 13-acetate was also effectively inhibited by these agents, but generally lower concentrations were required to obtain the same degrees of effects. These results indicated that some of the histamine H1 receptor-blocking drugs, such as clemastine and homochlorcyclizine, may act as inhibitors of formation of leukotrienes at the locus of inflammation.

A simple method for preparing mouse eosinophils was established, and the characteristics of the eosinophils were assessed including their responses to anti-allergic drugs.Mouse eosinophils were prepared from peritoneal exudate cells of BALB/c mice primed and boosted with antigen ovalbumin (OVA).Surface phenotypes, migration activities and leukotriene C(4) (LTC(4)) production abilities of these eosinophils were examined. In addition, the effects of anti-allergic drugs, oxatomide and tranilast, on generation of LTC(4) from mouse eosinophils were examined.Eosinophils of mice boosted with OVA were phenotypically and functionally identical with human eosinophils. Around 1 x 10(7) eosinophils were obtained from mouse peritoneal exudate. It was found that these mouse eosinophils enabled to migrate in response to eotaxin as well as platelet-activating factor (PAF), and generated LTC(4) by IL-5 stimulation. Moreover, it was revealed that clinically used anti-allergic drugs inhibited LTC(4)-production dose-dependently.The present study provides a convenient method to obtain fully functional mouse eosinophils that are useful for drug screening and for evaluating implications of eosinophils in allergic responses.

We report a 4-month-old female infant who developed West syndrome eleven days after administration of a histamine H1 antagonist, oxatomide, for atopic dermatitis. It has been reported that some histamine H1 antagonists induce seizures in epileptic patients. The age, the interval between oxatomide administration, and the onset of West syndrome and its clinical course were similar to two previously reported 3-month-old infants with West syndrome associated with ketotifen administration. We should be cautious in using the histamine H1 antagonists, oxatomide and ketotifen, in young infants because such agents could potentially disturb the anticonvulsive central histaminergic system.

The single peroral administration of the ethyl acetate extract or essential oil of German chamomile (Matricaria recutita L.) showed remarkable antipruritic effects in the compound 48/80-induced itch-scratching test in ddY mice, if suitable vehicle was used. The ethyl acetate extract or essential oil of German chamomile dissolved in the vehicle of 10% ethanol, 10% Tween 80 and 80% physiological saline was orally administrated 2 h before pruritus provocation by compound 48/80 subcutaneous injection. The ethyl acetate extract or essential oil of German chamomile showed significant dose-dependent inhibition of the compound 48/80-induced scratching without affecting spontaneous motor activity. The antipruritic effects of antihistamine H1 antagonists, oxatomide (10 mg/kg) and fexofenadine (10 mg/kg), were only partial in this test. However, the antipruritic effects of these agents were remarkably enhanced by the combined administration of the ethyl acetate extract of German chamomile (300 mg/kg). Thus, the co-medication with the ethyl acetate extract, or essential oil of German chamomile and antihistamines might be effective for the pruritus which could not be perfectly resolved alone by conventional antihistamines.

Oxatomide is an antiallergic drug used for the treatment of diseases mediated by type I allergy. Recently, terfenadine and astemizole, which have antiallergic actions similar to those of oxatomide, showed side effects on the cardiovascular system. This might be because concomitant drugs such as itraconazole inhibit cytochrome P450 3A4 (CYP3A4), the enzyme responsible for the degradation of terfenadine and astemizole, and thus the blood concentrations of the drugs are abnormally increased. In another article of this issue, we have reported that oxatomide is metabolized by CYP2D6-Val and CYP3A4, and simultaneously inhibits the metabolism of the model substrates for these enzymes. In this study, we performed the kinetic analysis of oxatomide metabolism using microsomes prepared from human liver, and found that the Km and Vmax values were 26.1 microM and 1254.4 pmol/mg protein/min, respectively. Ketoconazole, one of the representative inhibitors for CYP3A4, potently inhibited the metabolism of oxatomide, but other well-known CYP inhibitors did not show significant inhibition. These results suggest that the metabolism of oxatomide is principally catalyzed by CYP3A4. Furthermore, oxatomide inhibited the metabolism of (+/-) bufuralol and testosterone, model substrates for CYP2D6 and CYP3A4, respectively, in a dose-dependent manner with the Ki values of 57.4 and 24.3 microM, respectively. These observations, together with the finding that the putative highest concentration of oxatomide in blood was congruent with 40 ng/ml ( congruent with 93 nM) at 4 h after each dosage during consecutive 6-d administration, encouraged us to conclude that oxatomide won't inhibit CYP2D6 or CYP3A4 at clinical doses.

Oxatomide (CAS 60607-34-3) is an antiallergic agent effective against allergic rhinitis, urticaria, pruritus dermatitis, eczema dermatitis and bronchial asthma. The aim of this study was to establish the method for simultaneously determining oxatomide and its major metabolite M-11 in human serum, human plasma and rat plasma by high-performance liquid chromatography (HPLC). The method was applied to study the influences of alimentation on pharmacokinetics of oxatomide in rats. After extracting oxatomide and its metabolite M-11 from human serum, human plasma or rat plasma with diethyl ether under alkaline condition, sulfuric acid was added to the organic layer and oxatomide and M-11 were back-extracted. The aqueous layers were analysed by HPLC equipped with a fluorimetric detector. The method was highly sensitive and precise for quantitation of oxatomide and M-11 in human serum, human plasma and rat plasma in the concentration range of 1 to 125 ng/ml. Plasma concentration of oxatomide decreased biphasically with an elimination half-life (T1/2) of 1.59 h after intravenous administration of oxatomide (1 mg/kg) to non-fasting male rats. After oral administration of oxatomide (30 mg/kg) to fasting male rats, plasma concentration of oxatomide increased rapidly, and reached the maximum concentration of 188 ng/ml (Cmax) at 1.0 h. Plasma concentration of oxatomide decreased monophasically. The T1/2 was 2.58 h. The bioavailability was 6.74%. Plasma concentration of M-11 increased rapidly, and reached Cmax of 64.3 ng/ml at 0.7 h, and decreased monophasically with T1/2 of 3.79 h. After oral administration of oxatomide to non-fasting male rats, plasma concentration of oxatomide reached Cmax of 378 ng/ml at 3.3 h. The T1/2 was 3.27 h and the bioavailability was 17.5%. The Cmax and AUC0-infinity of M-11 were larger than those after oral administration of oxatomide to fasting male rats. These results demonstrated the usefulness of this method for monitoring and basic examination of biological samples and the influence of alimentation on absorption of oxatomide. Determination of plasma oxatomide concentrations would provide a useful indication of therapeutic efficacy.

The effects of Fujibitol, a preparation of crude drugs in wide clinical use for treatment of chronic rhinitis and empyema, on experimental allergic rhinitis in rats were studied. Fujibitol inhibited nasal allergic symptoms, i.e. sneezing and nasal rubbing, induced by antigen in sensitized animals. An increase in dye leakage into the nasal cavity induced by antigen was also inhibited by Fujibitol. On the other hand, no inhibitory effects were observed on either the nasal allergic symptoms or increase in dye leakage into the nasal cavity induced by histamine. However, Fujibitol was effective in inhibiting histamine release from the nasal cavity induced by antigen. Oxatomide used as positive control drug showed potent inhibitory effects on nasal symptoms and dye leakage into the nasal cavity induced by histamine and antigen. These results suggested that Fujibitol showed a remarkable protective effect against experimental rhinitis induced by antigen via inhibition of histamine release from the nasal cavity.

The effects of 4-benzhydryloxy-1-[3-(1H-tetrazol-5-yl)-propyl]piperidine (HQL-79), a newly developed antiallergic drug, on various chemical mediators and on chemical mediator release were investigated. Orally administered HQL-79 strongly inhibited the histamine-induced skin reaction in rats, and histamine- and 5-hydroxytryptamine (5-HT)-induced bronchoconstriction in guinea pigs. HQL-79 inhibited antigen-induced release of leukotriene (LT) B4, LTC4, histamine and prostaglandin (PG) D2 from the chopped lung tissues of actively sensitized guinea pigs. On the other hand, release of PGE2, one of the bronchoprotective prostanoids, was significantly enhanced by HQL-79. In an in vivo experiment, chronic administration of HQL-79 clearly reduced PGD2 contents and enhanced PGE2 contents in the lungs of repeatedly antigen-exposed guinea pigs. In biochemical studies, HQL-79 inhibited mouse spleen PGD synthase in a concentration-dependent manner. None of the antiallergics such as epinastine, terfenadine, oxatomide and cetirizine inhibited the PGD synthase. HQL-79 did not affect PGE synthase in sheep vesicular gland microsomes. These results suggest that antiallergic and antiasthmatic effects of HQL-79 could be ascribed to antihistaminic- and anti-5-HT effects, chemical mediator release inhibition, PGE2-release enhancement and PGD synthase inhibition. It is considered, in particular, that the differential modulation of PGD2 and PGE2 production is a conspicuous pharmacological feature of HQL-79.

The effects of histamine H 1 receptor antagonists on compound 48/80-induced scratching behavior were studied in mice. Classical histamine H 1 receptor antagonists such as diphenhydramine and chlorpheniramine caused a potent depressant effect on compound 48/80-induced scratching behavior. Histamine H 1 receptor antagonists having antiallergic activity (an inhibition of mast cell degranulation), such as azelastine and oxatomide and nonsedative histamine H 1 receptor antagonists such as terfenadine, epinastine and astemizole, also showed a relatively potent effect. On the other hand, the effects of tranilast and cromolyn sodium—antiallergic drugs without histamine H 1 receptor antagonistic activity—were extremely weak. Diazepam had weak or no depressant effects on compound 48/80-induced scratching behavior. These results suggest that inhibition of compound 48/80-induced scratching behavior is mainly due to histamine H 1 receptor antagonistic activity and not to the sedative action of the drugs.

To clarify the effects of anti-allergic drugs on substance P (SP) and vasoactive intestinal peptide (VIP) levels in nasal secretions, we employed competitive enzyme-linked immunoassays to measure concentrations of those neuropeptides in nasal secretions from 40 patients with house dust nasal allergy before and after administration of azelastine and oxatomide. One mg of azelastine and 30 mg of oxatomide were administrated twice a day for 4 weeks. Mean values of SP concentrations and ratios of SP to total protein of the nasal allergy group were significantly higher than those of the control group (p < 0.002). The VIP/total protein ratio of the allergy group was also significantly higher than that of the control group, although the VIP concentration alone was not. Mean levels of SP and VIP from patients with severe symptoms were significantly higher than those of the control group (p < 0.05), although those values were not significantly different between patients with moderate symptoms and control subjects. Azelastine and oxatomide effectively reduced SP levels in nasal secretions (p < 0.005), but they did not significantly decrease VIP levels. The reduction of SP levels was significant in patients with excellent responses to those drugs (p < 0.005), but not in patients with poor responses. These findings suggest that SP and VIP levels in nasal secretions may reflect the clinical state of nasal allergy and be one of the better parameters available for evaluating the clinical efficacy of anti-allergic drugs against nasal allergy.

In a mast cell model, oxatomide displays inhibition of mediator release which is not related to its histamine H1 receptor antagonistic activity. From a previous study it appeared that especially early steps in the signal transduction leading to exocytosis were influenced by oxatomide. We now studied effects of oxatomide on those early steps in more detail. The antigen- and thapsigargin-mediated exocytosis in rat basophilic leukemia (RBL-2H3) cells were both inhibited by oxatomide. After aggregation of high affinity receptors for immunoglobulin E (Fc epsilon RI), protein tyrosine phosphorylation is induced. Oxatomide caused remarkable changes in the tyrosine phosphorylation pattern in resting cells. Also after antigen and thapsigargin activation, changes in the tyrosine phosphorylation of cellular proteins are observed. In addition, Ca2+ fluxes were studied by means of the net influx of 45Ca2+ and by measuring intracellular free Ca2+ concentrations ([Ca2+]) with the fluorescent probe fura-2. Oxatomide inhibited the 45Ca2+ influx and the increase in [Ca2+]i upon antigen and thapsigargin activation of the cells. Neither the release of Ca2+ from internal stores nor the efflux of Ca2+ over the plasma membrane seems to be affected. The effect of oxatomide on Ca2+ influx was further characterized by studying Ba2+ influx in the absence of extracellular free Ca2+. We conclude that inhibition of mediator release is mainly caused by inhibition of influx of extracellular Ca2+, via plasma membrane Ca2+ channels that are activated by depletion of intracellular Ca2+ stores. The molecular mechanism with which oxatomide might interfere with these channels is discussed.

Background Azelastine, oxatomide, and ketotifen are used for patients with allergic diseases. These drugs inhibit the release of chemical mediators including the Leukocyteendothelial; however, the mechanism involved is unclear. Objective To clarify the mechanism of inhibition, we investigated the effects of three drugs on the function of phospholipase A 2 , 5-lipoxygenase, leukotriene C 4 synthase, and leukotriene A 4 hydrolase, which are all catabolic enzymes involved in synthesizing leukotriene C 4 and leukotriene B 4 in rat basophilic leukemia (RBL)-1 cells. Methods and results The production of leukotriene C 4 and leukotriene B 4 was measured by high performance liquid chromatography (HPLC). All three drugs inhibited the production of leukotriene C 4 and leukotriene B 4 when cells were stimulated with A23187. All three drugs also inhibited the A23187-stimulated release of 3 Harachidonic acid from membrane phospholipids. Azelastine inhibited the production of leukotriene C 4 , but not leukotriene B 4 , when either arachidonic acid or leukotriene A 4 free acid was used as the substrate in our cell free system. Oxatomide and ketotifen did not inhibit the synthesis of either leukotriene C 4 or leukotriene B 4 in the same cell free study. Conclusion Results indicated that oxatomide and ketotifen inhibit the production of leukotriene C 4 and leukotriene B 4 by inhibiting phospholipase A 2 activity, whereas, azelastine inhibits the leukotriene C 4 production by inhibiting phospholipase A 2 and leukotriene C 4 synthase.

We investigated the effect of topically applied oxatomide, an antiallergic agent, on the assault of allergic rhinitis in actively sensitized guinea pigs. Topical application of oxatomide nasal spray (0.025%) reduced the severity of allergic rhinitis which was assessed by determining dye leakage and histamine released to the nasal cavity of guinea pigs. Furthermore, oxatomide nasal spray treatment significantly prevented the increase in dye leakage induced by histamine administration in guinea pigs and rats. These results indicate that the topical application of oxatomide inhibits both the release and the action of histamine. Therefore, oxatomide nasal spray may be beneficial for treatment of allergic rhinitis.

In a model for mucosal mast cells (RBL-2H3 cells) a set H1-antagonist derived anti-allergic drugs containing a diphenylmethyl piperazinyl moiety was examined for their ability to inhibit release of the mediator beta-hexosaminidase. Cells were activated with antigen or the calcium ionophore A23187, whether or not in combination with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA). Oxatomide, hydroxyzine and cetirizine inhibit the antigen induced beta-hexosaminidase release. The release triggered by A23187, whether or not in combination with TPA is hardly influenced by the compounds. A biphasic dependence of the inhibition of exocytosis in RBL cells on lipophilicity is observed with the optimum at log P is 5-6. The extremely lipophilic compounds meclozine and buclizine are not active in this model. pH dependence of the effect of the drugs shows that especially the uncharged species are active in inhibiting exocytosis. The investigated compounds show an effect on phase transitions in L-alpha-phosphatidylcholine dipalmitoyl liposomes as assayed with differential scanning calorimetry (DSC). For the less extremely lipophilic compounds the induced changes in the phospholipid membranes increased with lipophilicity. The relation between structural features of the drug and the interaction with phospholipids is discussed in view of the DSC results. We conclude that location of the active drugs at the membrane or the membrane/protein interface is important for the inhibiting activity on exocytosis. This could affect several membrane related processes, which are abundant in the early phases of the IgE-mediated signal transduction process.

The present study was performed to investigate the putative suppressive effects of H1-receptor antagonists (HRA) of the second generation (astemizole (AS), cetirizine (CT), loratadine (LO), oxatomide (OX) and terfenadine (TF)) on the mediator release from human basophils activated by two classical stimuli. Anti-IgE-mediated histamine release was inhibited in a dose-dependent fashion by TF (maximum inhibitory value: 33.8 +/- 7.6%, 100 microM, n = 7), whereas the other HRA exhibited weaker activity. The anti-IgE-induced LTC4 production was strongly suppressed by TF, LO and OX (92.4 +/- 6.3%, 90.8 +/- 6.0% and 88.5 +/- 5.6%, 100 microM, n = 4-5), while AS was less active (56.4 +/- 4.1%, 100 microM, n = 5). Histamine release induced by incubation with grass pollen antigen (0.01%) was inhibited by TF (40.7 +/- 4.1%, 50 microM, n = 4), but the other HRA showed only low activity. The present findings suggest that some HRA might exhibit direct inhibitory effects on activation of IgE-receptor bearing cells.

This study was designed to analyse prostaglandins (PGs) in human nasal lavage fluid using the combination of microcolumn high performance liquid chromatography and a He/Cd laser induced fluorescence detection system. Forty-seven patients with allergic rhinitis and 12 healthy volunteers were investigated. Four species of PG, i.e. PGD2, PGE2, PGF2 alpha, and 6-keto-PGF1 alpha were detected in the nasal lavage fluid. Concentrations of PGD2 (1.33 +/- 0.17 nmol/ml) and PGE2 (0.87 +/- 0.11 nmol/ml) in nasal lavage fluid from patients with allergic rhinitis (the allergy group) were significantly increased compared with those of volunteers (the control group, 0.23 +/- 0.16 nmol/ml, 0.29 +/- 0.19 nmol/ml, respectively). On the other hand, no significant differences were observed in concentrations of either PGF2 alpha or 6-keto-PGF1 alpha between the control group and the allergy group. Histamine concentration in nasal lavage fluid was significantly increased in the allergy group (53 +/- 7.6 nmol/l) compared with the control group (3.4 +/- 1.0 nmol/l). No significant correlation was observed between PGD2 and histamine concentration (r = 0.24), or between PGE2 and histamine concentration (r = 0.08) in nasal lavage fluid from patients with allergic rhinitis. Treatment with oxatomide, an anti-histamine and anti-allergic drug, significantly improved symptom scores, but did not alleviate them completely. Concentrations of each PG detected in nasal lavage fluid did not change significantly after oxatomide treatment. It is concluded that, not only histamine but also PGs, particularly PGD2 and PGE2, might be involved in the genesis of allergic rhinitis.

The effect of oxatomide on reactive oxygen species (ROS) generated both by neutrophils and in a cell-free, xanthine-xanthine oxidase system was examined. The species investigated were superoxide radical anion (O2-), hydrogen peroxide (H2O2) and hydroxyl radical (OH.). Oxatomide significantly decreased neutrophil-generated O2-, H2O2 and OH. in a dose-dependent manner. H2O2 and OH. generated in a cell-free system were also reduced in the presence of oxatomide. The present study indicates that oxatomide decreases ROS generation presumably by inhibiting the neutrophil oxygen metabolism, and has the ability to scavenge H2O2 and OH.

The effects of anti-allergic drugs with or without H1-receptor antagonism on the NADPH oxidase (EC 1.6.99.6) from human neutrophils in both whole-cell and fully soluble (cell-free) systems were investigated. Three anti-allergic drugs with H1-receptor antagonism, azelastine, ketotifen and oxatomide, were found to inhibit the superoxide generation of human neutrophils exposed to phorbol myristate acetate in a whole-cell system and the activation of superoxide-generating NADPH oxidase by sodium dodecyl sulfate in a cell-free system. The concentrations of these drugs required for 50% inhibition of the oxidase (IC50) were: azelastine--0.7 microM in the whole-cell system and 0.5 microM in the cell-free system; ketotifen--60 microM in the whole-cell system and 6.8 microM in the cell-free system; and oxatomide--25 microM in the whole-cell system and 9.7 microM in the cell-free system. In addition, in the cell-free system, these drugs did not change the Km values for the NADPH of the oxidase. However, these drugs did not inhibit the superoxide generation of NADPH oxidase after its activation in whole-cell and cell-free systems, suggesting that these drugs do not have superoxide-scavenger actions. Concentrations of less than 200 microM of anti-allergic drugs without H1-receptor antagonism, tranilast, repirinast and ibudilast, did not inhibit neutrophil NADPH oxidase in whole-cell and cell-free systems. The IC50 of hydrocortisone in the cell-free system was 60 microM. These results suggest that anti-allergic drugs with H1-receptor antagonism inhibit activation of the solubilized membrane-bound enzyme by sodium dodecyl sulfate in cell-free systems and that they have much stronger anti-inflammatory action than hydrocortisone.

Ro 21-7634 was examined for oral antiallergic activity in two in vivo models commonly used to evaluate antiallergics. In the rat PCA test, this drug had an oral ID50 of 1.14 mg/kg and was found to be more potent than several other antiallergics including Disodium Cromoglycate (cromoglycate), Oxatomide, Doxanthrazole, Xanoxate, 2,6-bis (ethyoxyoxalylamino) pyridine, PRD-92-EA and M + B 22,948. In contrast to cromoglycate, Ro 21-7634 was found to be an orally active inhibitor of antigen-induced broncho-constriction in passively sensitized rats (ID50 = 0.2 mg/kg). In addition, Ro 21-7634 inhibited antigen-induced histamine release in an in vivo passive peritoneal anaphylaxis test system, following intraperitoneal administration. Ro 21-7634 demonstrated no end organ antagonism toward histamine, metacholine or serotonin in the guinea pig.

A new series of 3-(3-pyridyl)acrylamides 16, 17, 19, and 26, and 5-(3-pyridyl)-2,4-pentadienamides 20-25 were prepared and evaluated for their antiallergic activity. Several of these compounds exhibited more potent inhibitory activities than the parent compound 1a [(E)-N-[4-[4-(diphenylmethyl)-1-piperazinyl]butyl]-3- (3-pyridyl)acrylamide] against the rat passive cutaneous anaphylaxis (PCA) reaction and the enzyme 5-lipoxygenase. Particularly, (E)-N-[4-[4-(diphenylmethyl)-1-piperazinyl]butyl]-3- (6-methyl-3-pyridyl)acrylamide (17p) showed an ED50 value of 3.3 mg/kg po in the rat PCA test, which was one-fifth of ketotifen and oxatomide. As compared with ketotifen and oxatomide, compound 17p (AL-3264) possessed a better balance of antiallergic properties due to inhibition of chemical mediator release, inhibition of 5-lipoxygenase, and antagonism of histamine.

We have studied the ability of four H1 antihistamines, ketotifen, oxatomide, clemastine and promethazine, and of disodium cromoglycate (DSCG) to inhibit in vitro histamine release mediated

The ultrastructure of isolated rat peritoneal mast cells was evaluated after in vitro degranulation and treatment with oxatomide, a new anti-allergic compound. In a first series of experiments, mast cells of rats infected with Trichinella spiralis larvae were incubated with Trichinella larvae somatic antigen to produce histamine release. The release was visualized in the electron microscope by exocytosis of the peripheral amine-containing granules, which resulted from fusion between several perigranular membranes and fusion of these membranes with the plasma membrane. A more drastic degranulation was provoked in a second series by incubation of unsensitized mast cells in the presence of the amine liberator compound 48/80. This treatment led to a complete extrusion of the granules in most of the cells, while in a smaller number of cells, only large vacuoles containing remnants of several granules were seen. The plasma membrane of these cells however was intact and there were no signs of exocytosis. The effect of oxatomide against mast cell degranulation was dose-dependent and comparable for the two types of histamine release. After incubation with high doses (10(-4) M, 5.10(-5) M) granule liberation was rarely observed in antigen-challenged and compound 48/80-challenged cells. Protection was apparently situated at the level of the plasma membrane which seemed to be unable to fuse with the perigranular membranes while fusion of perigranular membranes of individual granules was still possible. None of the tested concentrations of oxatomide induced spontaneous degranulation. High doses, however, led in a number of cells to some ultrastructural alterations such as partial disappearance of plasmalemmal folds, slight cytoplasmic oedema and the appearance of intranuclear microtubules. The latter were also seen in oxatomide-treated challenged mast cells.

Oxatomide is a very potent inhibitor of histamine release induced by anti-1gE and Timothy pollen extract in passively sensitized human lung fragments and that induced by anti-1gE from human leucocytes. Its spectrum of activity is different from sodium cromoglycate-like drugs. In high concentrations oxatomide, like other antihistamines and related structures, induced histamine release from both lung and leucocytes. However, oxatomide induced histamine release far more effectively from sensitized lung than non-sensitized lung. Inhibition of immunologically induced histamine releast by oxatomide may play a part in its action as an anti-hay fever and, possibly, an anti-asthmatic drug.

11-Chloro-6,11-dihydrodibenzo[b,e]thiepin and its 2-methyl derivative VI were transformed via the 11-(4-ethoxycarbonyl)-1-piperazinyl) compounds IVc and Vc to 11-(1-piperazinyl)compounds IVb and Vb. Their reactions with 1-(3-chloropropyl)-1,3-dihydro-2H-benzimidazol-2-one (II) afforded the title compounds IVa and IVb. Similar reactions and sequences in the series of 10,11-dihydro-5H-dibenzo[a,d]cycloheptene, 4,9-dihydrothieno[2,3-c]-2-benzothiepin, and 10,11-dihydrodibenzo[b,f]thiepin led to further oxatomide (Ia) analogues IIIa and VIIa-XIa. In the test of passive cutaneous anaphylaxis in rats, compound VIIIa was more active than Ia, and IVa had similar activity like Ia.

The role of histamine in the cardiovascular responses to hypoxia has not been resolved. Oxatomide, an orally active anti-allergic drug, has been reported to inhibit mast cell discharge and to antagonize any histamine which is released. This drug was administered to six dogs to determine the effect of inhibiting histamine release on the pulmonary vascular and cardiac responses to hypoxia. Oxatomide potentiated hypoxic pulmonary vasoconstriction, but reduced the cardiac output and stroke volume responses to hypoxia. These findings suggest that histamine is released during hypoxia and acts to oppose hypoxic vasoconstriction. In addition, histamine released from peripheral tissues may play an important role in mediating the increased cardiac output observed during acute hypoxia. By utilizing infusions of histamine, oxatomide was also found to be a histamine H1-receptor antagonist.

Plasma levels, biotransformation and excretion of oxatomide were studied after single oral doses of 14C-oxatomide in male rats, dogs and humans. Oxatomide was very well absorbed, and almost completely metabolized in the three species. Excretion of the metabolites was very rapid and complete within a few days; the 14C label was excreted more in the faeces (54-62%) than in the urine (27-40%). Major metabolic pathways of oxatomide were oxidative N-dealkylations at the piperazine nitrogens and at the benzimidazolone nitrogen in rats and man, and also aromatic hydroxylation at the benzimidazolone moiety in man. The main urinary metabolite in the three species was 2,3-dihydro-2-oxo-1H-benzimidazole-1-propanoic acid, resulting from the oxidative N-dealkylation at the 1-piperazine nitrogen.

Different types of anti-allergic agents have been compared for their ability to inhibit the antigen-induced contraction of isolated lung parenchymal strips and ilea of guinea pigs. Extremely low concentrations of the lipoxygenase inhibitor NDGA inhibit the contraction of the lung parenchymal strips and higher concentrations block the ileal phase II-contraction (both predominantly caused by the developing SRS-A), but do not inhibit ileal phase I-contraction (caused by released histamine). The H1-antihistamine clemastine predominantly antagonizes phase I of the ileum contraction, and the anti-allergic drug oxatomide reduces all three types of contraction. The degranulation inhibitor DSCG does not influence the contraction of the lung strip, but produces a slight inhibition of both ileal contractions. The PAF-antagonist BN 52021 influences these three contractions in the opposite way to DSCG. As these various types of anti-allergic agents influence these three contractions in a different manner, the models can be used for judging the mechanism of action of a new anti-allergic compound.

SUMMARY The antagonistic activity of oxatomide, and its effects on evoked histamine release and histamine-N-methyl transferase activity in skin, have been studied. Oxatomide antagonizes HI activity in a dose-dependent but non-competitive manner. It also shows seme atropine-like activity. Oxatomide did not cause detectable inhibition of antigen-stimulated histamine release from skin slices of sensitized guinea-pigs although the possibility that oxatomide may cause weak inhibition could not be excluded. In the presence of low concentrations of histamine, oxatomide suppressed human skin histamine-N-methyl transferase, but in the presence of higher substrate concentrations it enhanced activity of this enzyme. These observations; which were limited by the poor solubility of oxatomide in aqueous media, should encourage further in vivo studies of oxatomide's histamine-suppressing properties in the human subject.

Several Ca2+ antagonists with either Ca2+-entry blocking or calmodulin (CaM) antagonistic properties and antiallergic drugs were investigated for their effects on mediator release from mast cells induced by different secretagogues (compound 48/80, concanavalin A, antigen-IgE and Ca2+ ionophore A23187) and for their ability to inhibit the function of CaM or phospholipid/Ca2+-dependent protein kinase (C-kinase). The effects of the different agents--with the only exception of cromolyn sodium--on histamine release elicited by compound 48/80 correlated well with their actions on two CaM-dependent enzymes whereas the activity of C-kinase was far less altered, or not altered at all. CaM antagonism of cloxacepride, picumast, oxatomide, fendiline and bepridil correlated not only with the inhibition of exocytosis evoked by compound 48/80 but also with that induced by A23187, concanavalin A and antigen-IgE. This indicates an action of these substances distal to the generation of the Ca2+ signal since the various secretagogues elevate the intracellular Ca2+ concentration by different mechanisms. However, prenylamine and thioridazine inhibited concanavalin A- and antigen-IgE-induced mediator release more potently and more effectively than that elicited by compound 48/80 or A23187. Therefore inhibition of allergic histamine release by these drugs may in part be dependent on an impairment of the Ca2+ signal. Since for each of two agents inhibition of histamine release (evoked by different releasers) parallels that of serotonin release it may be concluded that these mediators are secreted via the same mechanism. The results obtained with agents exhibiting different pharmacological properties but which share one common property, namely antagonism of CaM, strengthen the view that CaM is involved in exocytosis of mediators from mast cells.