In Vitro and in Vivo Activity Profile of RHC 2851: A New Orally Effective Antiallergic Agent

RHC 2851 has been investigated for its antiallergic activity in three in vitro and two in vivo models of anaphylaxis. We have also compared its activity profile in these models with that of disodium cromoglycate (DSCG), doxantrazole, ketotifen and oxatomide. RHC 2851, given i.p., was 6 times more potent than DSCG, and given orally it was 3 times more potent than doxantrazole. As an inhibitor of mediator release, the activity profile of RHC 2851 was identical to that of DSCG in the following respects: inhibition of IgE-mediated in vitro release of histamine from rat mast cells (RMC) but not human basophils (HUB), possession of tachyphylactic properties and demonstration of rapid loss of inhibitory activity as a function of time before antigen challenge as well as inability to inhibit IgG₁-mediated release of histamine, both in vitro and in vivo, and lack of mediator antagonist activity. Ketotifen and oxatomide did not inhibit either IgE or IgG₁-mediated release of histamine in vitro or in vivo, and were potent mediator antagonists in vivo. We conclude that RHC 2851 is an orally effective inhibitor of mediator release with a mechanism of action similar to that of DSCG and different from that of ketotifen and oxatomide.     

HIV-1 infection complicated by food allergy and allergic gastroenteritis: a case report

A 34-year-old female with HIV-1 infection detected by positive serology in 1983 subsequently developed acute granulomatous interstitial pneumonitis, eosinophilic gastroenteritis, and angioedema associated with the ingestion of vanilla ice cream and tangerines. The enteritis and angioedema symptoms appeared to respond to large doses of oral sodium cromoglycate. Sera collected over several years before clinical symptoms revealed a sharp rise of IgE antibody in 1985 and a subsequent decline to baseline values followed by markedly increased levels of IgE antibodies to a number of inhalant and food allergens. The findings suggest disordered IgE antibody regulation as a consequence of HIV-1 infection and as a cause of allergic manifestations including eosinophilic gastroenteritis and food-induced angioedema.     

Inhibition by azelastine of nonallergic histamine release from rat peritoneal mast cells

The ability of azelastine and selected antiallergic drugs to inhibit compound 48/80-induced and PS-potentiated, Con A-induced histamine release from RPMC was investigated. Azelastine, ketotifen, theophylline, and DSCG added simultaneously with the secretagogues or preincubated with the RPMC for 10 min before the addition of secretagogues produced concentration-dependent inhibition of histamine release. In general, the relative order of potency at calculated IC50 level was as follows: azelastine greater than ketotifen greater than theophylline greater than DSCG. The preincubation of RPMC with azelastine for 10 min exerted 3.5 times greater inhibition of Con A plus PS-stimulated histamine release but did not influence the inhibitory activity on compound 48/80-induced release. The duration of preincubation did not influence the inhibitory effects of ketotifen with either secretagogue. Theophylline and DSCG exerted significantly greater inhibition when they were added simultaneously with Con A plus PS. The inhibitory activity of DSCG was also significantly improved upon simultaneous addition with compound 48/80. These data demonstrated that azelastine is the most potent inhibitor of nonallergic histamine release from RPMC among the four antiallergic drugs examined.     

Inhibition of allergic histamine release by azelastine and selected antiallergic drugs from rabbit leukocytes

The ability of azelastine to inhibit allergic histamine release from rabbit mixed leukocytes was studied and compared with selected antiallergic drugs. Azelastine, ketotifen, diphenhydramine, theophylline and disodium cromoglycate (DSCG) produced concentration-dependent inhibition of allergic histamine release from rabbit basophils. The concentrations inhibiting histamine release by 50% (IC50; microM) were as follows: azelastine = 4.5; ketotifen = 9.5; diphenhydramine = 18.9; theophylline = 56.9; DSCG = greater than 1,000. DSCG was added to the cells immediately prior to antigen challenge. All other drugs were preincubated for a period of 10 min prior to antigen challenge. At the IC50 level, azelastine is about 2, 4, 13 and greater than 200 times as effective as ketotifen, diphenhydramine, theophylline and DSCG, respectively. The IC50 of azelastine following 0, 10 and 30 min preincubation were 2.4, 1.9 and 3.5 microM, respectively. These observations showed: (1) azelastine is capable of acting rapidly on basophils and of inhibiting allergic histamine secretion, and (2) the prolongation of the preincubation time of azelastine up to 30 min with rabbit leukocytes did not exhibit any sign of tachyphylaxis (loss of activity). In conclusion, azelastine is a potent inhibitor of allergic histamine secretion from the leukocytes of ragweed-sensitized rabbits.     

In Vitro and in Vivo Activity Profile of RHC 2851: A New Orally Effective Antiallergic Agent

Abstract

RHC 2851 has been investigated for its antiallergic activity in three in vitro and two in vivo models of anaphylaxis. We have also compared its activity profile in these models with that of disodium cromoglycate (DSCG), doxantrazole, ketotifen and oxatomide. RHC 2851, given i.p., was 6 times more potent than DSCG, and given orally it was 3 times more potent than doxantrazole. As an inhibitor of mediator release, the activity profile of RHC 2851 was identical to that of DSCG in the following respects: inhibition of IgE-mediated in vitro release of histamine from rat mast cells (RMC) but not human basophils (HUB), possession of tachyphylactic properties and demonstration of rapid loss of inhibitory activity as a function of time before antigen challenge as well as inability to inhibit IgG₁-mediated release of histamine, both in vitro and in vivo, and lack of mediator antagonist activity. Ketotifen and oxatomide did not inhibit either IgE or IgG₁-mediated release of histamine in vitro or in vivo, and were potent mediator antagonists in vivo. We conclude that RHC 2851 is an orally effective inhibitor of mediator release with a mechanism of action similar to that of DSCG and different from that of ketotifen and oxatomide.     

Allergen-specific immunotherapy with a monophosphoryl lipid A-adjuvanted vaccine: reduced seasonally boosted immunoglobulin E production and inhibition of basophil histamine release by therapy-induced blocking antibodies

BACKGROUND: Allergen-specific immunotherapy represents a causal form of treatment for IgE-mediated allergies. The allergen extract-based analyses of immunotherapy-induced effects yielded highly controversial results regarding a beneficial role of therapy-induced IgG antibodies. OBJECTIVE: We analysed allergen-specific IgE, IgG subclass, and IgM responses in patients treated with a grass pollen allergy vaccine adjuvanted with monophosphoryl lipid A (MPL), a Th1-inducing agent, and in a placebo group using recombinant timothy grass pollen allergen molecules (rPhl p 1, rPhl p 2, rPhl p 5). RESULTS: The strong induction of allergen-specific IgG1 and IgG4 antibodies observed only in the actively treated group was associated with significant clinical improvement. Therapy-induced allergen-specific IgM and IgG2 responses were also noted in several actively treated patients. An inhibition of allergen-dependent basophil histamine release was only obtained with sera containing therapy-induced allergen-specific IgG, but not with sera obtained before therapy or from placebo-treated patients. Moreover, patients with therapy-induced allergen-specific IgG antibodies showed a reduced induction of allergen-specific IgE responses during seasonal grass pollen exposure. CONCLUSION: Successful immunotherapy with the MPL-adjuvanted grass pollen allergy vaccine is associated with the production of allergen-specific IgG antibodies. These blocking antibodies may have protective effects by inhibiting immediate-type reactions and systemic increases of IgE responses caused by seasonal allergen exposure.     

IgE antibodies against bovine serum albumin in a case of eosinophilic gastroenteritis

Eosinophilic gastroenteritis is a disease characterized histologically by an eosinophilic infiltration of the gut. The cause of this disease remains unclear, although both food allergy and food intolerance have been implicated in its pathogenesis. We report the case of a 22-year-old man in whom gastrointestinal symptoms first appeared in childhood, with involvement of mucosa and muscularis layers of stomach and bowel. He presented high IgE blood levels, and his prick test was positive to bovine, pig, and lamb sera. Immunoblots from calf, pig, and lamb sera, incubated with the patient's serum and revealed by autoradiography, demonstrated the presence of a 65-kDa protein band that was recognized by IgE antibodies but not by IgG. This band corresponded to bovine serum albumin, while IgE did not show reactivity with human albumin. These data suggest a possible role for IgE-mediated hypersensitivity mechanisms in the pathogenesis of eosinophilic gastroenteritis.     

Saturation of immunoglobulin E (IgE) binding sites by polyclonal IgE does not explain the protective effect of helminth infections against atopy

One hypothesis for the decreased rates of atopy observed among helminth-infected individuals is that parasite-induced polyclonal immunoglobulin E (IgE) out-competes allergen-specific IgE for FcepsilonRI binding on basophils and mast cells. In experiments with fresh blood drawn from filaria-infected patients, we found no association between ratios of polyclonal to Brugia malayi antigen (BmAg)-specific IgE (range, 14:1 to 388:1) and basophil responses to BmAg as measured by histamine release. Using serum samples from a filaria-infected patient who also had dust mite (Dermatophagoides pteronyssinus)-specific IgE antibodies from time points with various ratios of polyclonal to D. pteronyssinus-specific IgE (16:1 to 86:1), we demonstrated that increased ratios of polyclonal to D. pteronyssinus-specific IgE did not attenuate basophil sensitization as measured by D. pteronyssinus-specific histamine release. Suppression of histamine release was likely not observed in either of these sets of experiments because polyclonal to antigen-specific IgE ratios were not sufficiently high, as concurrent passive sensitization of basophil experiments required ratios of polyclonal to antigen-specific IgE of greater than 500:1 to suppress basophil histamine release. Further, the intensity of IgE staining in basophil populations from 20 patients with active filaria infections correlated strongly with total serum IgE levels (rho = 0.698; P = 0.0024) with no plateau in intensity of IgE staining, even though some patients had total IgE levels of greater than 10,000 ng/ml. Our data therefore suggest that in helminth infections (and in filarial infections in particular), the ratios of polyclonal to allergen-specific IgE rarely reach those levels necessary to inhibit allergen-specific IgE-FcepsilonRI binding and to suppress allergen-induced degranulation of mast cells and basophils.     

Allergy to cow's milk with onset in adult life

We report a 29-year-old patient with a history of asthma that is sometimes accompanied by urticaria, which is related to the ingestion of milk products. Once these were excluded from his diet no symptoms were observed. Skin tests, specific IgE, histamine release, and oral food challenges were positive to cow's milk and its fractions.     

The stripped basophil histamine release bioassay as a tool for the detection of allergen-specific IgE in serum

BACKGROUND: For the detection of allergen-specific IgE in serum, IgE-binding assays such as the radioallergosorbent assay (RAST) are commonly used. In this study, the applicability and sensitivity of the stripped basophil histamine release bioassay was investigated and compared to the RAST. METHODS: Basophils were stripped of their IgE by an acidic buffer, sensitized by human serum and stimulated by allergen with or without interleukin (IL)-3. The histamine release was determined by fluorometric analysis. RESULTS: We showed that for enhancement of the maximal histamine release and the sensitivity of the stripped basophil assay, the priming cytokine IL-3 can be added to the basophils simultaneously to the stimulus. Preincubation of the cells with IL-3, as described in other studies, was not necessary. The bioassay can be used to study the specificity of IgE-mediated reactions. Basophils sensitized by serum absorbed to a particular allergen did not respond to this allergen anymore. This method is very suitable to study cross-reactivity between allergens. The results obtained in the bioassay were comparable to those obtained in the RAST. Using the RAST, lower concentrations of allergen-specific IgE were detected than in the bioassay. However, sera containing IgE against minor allergenic components were negative in the RAST, but strongly positive in the basophil assay. CONCLUSIONS: The stripped basophil histamine release bioassay is useful to complement and extend serological detection of allergen-specific IgE. Especially with sera containing IgE against minor components, this assay is more suitable than the RAST. Furthermore, in this assay, the dependency of IgE and of allergen-specific IgE in reactions can be studied in more detail.     

Long-Term Modification on Histamine-Induced Bronchoconstriction by Disodium Cromoglycate and Ketotifen versus Placebo

In order to compare long-term changes induced on a wide spectrum of bronchial hyperreactivity (BH) by the antiallergic drugs disodium cromoglycate (DSCG) and ketotifen, 56 asthmatic patients (age range 15-55 years) were studied. Patients were allocated to three groups with similar age and BH level. During 2 months, 15 individuals inhaled 20 mg DSCG four times a day, 14 took ketotifen 1 mg twice a day orally and 14 one placebo (lactose) capsule twice a day. After preliminary results, an additional group of 13 patients took clemastine 1 mg orally twice a day for 1 week. Only the ketotifen and clemastine groups differed significantly from the placebo group on shifting log dose-response curves of inhaled histamine. In addition, no significant difference was seen between the ketotifen and clemastine groups. These results suggest that changes induced by ketotifen were mainly related to its powerful antihistamine action; similarly, nonspecific BH is not wholly dependent on mediator release.     

Actions and cross-reactivity of antiallergic agents and a calcium channel antagonist on rat peritoneal mast cells. Difference in the action mechanisms and cross-reactivity among the agents

The actions of the antiallergic agents, disodium chromoglycate (DSCG), tranilast and ketotifen, and of a calcium channel antagonist, nicardipine, and cross-reactivity among the agents were examined by observing the inhibition of⁴⁵Ca uptake and histamine release in rat mast cells stimulated by antigen and compound 48/80 (comp. 48/80).

1. All agents inhibited⁴⁵Ca uptake and histamine release in mast cells stimulated by antigen. The inhibition of⁴⁵Ca uptake by the antiallergic agents paralleled the inhibition of histamine release, while nicardipine inhibition of⁴⁵Ca uptake was stronger than its inhibition of histamine release.
2. The action of DSCG on⁴⁵Ca uptake and histamine release was significantly decreased in cells stimulated with antigen and phosphatidylserine (PS), while tranilast inhibition of histamine release was not affected by the addition of PS despite a significant decrease in the inhibition of⁴⁵Ca uptake.
3. The inhibitory effect of DSCG and tranilast was significantly lower in mast cells stimulated by comp. 48/80 than in the cells stimulated by antigen.
4. Tachyphylaxis was observed in cells re-exposed to DSCG and tranilast following previous exposure to the agents.
5. Cross-reactivity was found between DSCG and tranilast.

     

Spontaneous release of histamine from basophils and histamine-releasing factor in patients with atopic dermatitis and food hypersensitivity

Patients with hypersensitivity to food documented by a double-blind, placebo-controlled oral food challenge have been reported to have a high rate of release of histamine from basophils in vitro. To determine whether patients with atopic dermatitis and food hypersensitivity had similar high rates of spontaneous histamine release in vitro, whether dietary elimination of relevant food antigens affected this release, and whether a cytokine, histamine-releasing factor, could account for it, we evaluated 63 patients with atopic dermatitis and food hypersensitivity (38 of whom had eliminated the offending foods from their diets), 20 patients with atopic dermatitis without food hypersensitivity, and 18 normal volunteers. Patients with atopic dermatitis and food hypersensitivity were found to have higher rates of spontaneous release of histamine from basophils than controls (mean +/- SE, 35.1 +/- 3.9 percent vs. 2.3 +/- 0.2 percent; P less than 0.001). Those who had eliminated the offending food allergen from the diet for an extended period had a significantly lower rate of histamine release (3.7 +/- 0.5 percent; P less than 0.001). In patients with atopic dermatitis without food hypersensitivity, the rate (1.8 +/- 0.2 percent) did not differ from that in normal controls. Mononuclear cells from persons with food allergies spontaneously produced a histamine-releasing factor in vitro that provoked the release of histamine from the basophils of other food-sensitive persons, but not from those of normal controls. Patients who adhered to a restricted diet had a decline in the rate of spontaneous generation of the factor by their mononuclear cells. The histamine-releasing factor was found to activate basophils through surface-bound IgE. We conclude that in patients with food hypersensitivity, exposure to the relevant antigens produces a cytokine (histamine-releasing factor) that interacts with IgE bound to the surface of basophils, causing them to release histamine.     

Antigen-induced histamine-release from duodenal biopsy in gastrointestinal food allergy

Twenty-four patients allergic to food, which was demonstrated by oral provocation, were investigated. Six particles from duodenal mucosa were obtained during endoscopic examination and incubated with different foods. Specimens challenged by anti-human-IgE or without any stimulus served as control values. Also, skin tests and assays of specific IgE were performed. The spontaneous histamine release varied from 19% to 36%. Anti-IgE caused an increase up to 26% until 65%. Incubation with allergenic food induced a histamine release from 41% to 81%, demonstrating positive results in 27 out of 30 separate experiments. Antigen-induced histamine release from biopsy specimens turned out superior to skin tests and specific IgE. It is the most reliable tool for diagnosis of gastrointestinal food allergy besides oral provocation.     

In vitro assays for the diagnosis of IgE-mediated disorders

Advances in technology have provided new laboratory tools for the quantitation of allergen-specific IgE antibodies in serum and on the surface of basophils. This review examines the evolution from qualitative IgE antibody assays of the late 1960s to the present-day, third-generation, automated and quantitative allergen-specific IgE assays. The latest technology trend is toward microarrays in which crude or purified native and recombinant allergens can be spotted in microdot arrays on silica chips to permit extensive panels of specific IgE measurements to be performed with small quantities of serum. Although these technologies hold promise, their diagnostic performance requires further assessment once their technical details have been optimized. Potential abuses of this newer IgE antibody technology include the use of allergosorbent specificities (eg, especially food and drugs) that lack validation, application of IgE antibody measurements in the diagnosis of non-IgE-dependent disorders (eg, aspirin sensitivity), and modification of IgE antibody assays to measure food-specific IgG antibody for which there is no clinical indication. Basophil mediator release assays have evolved to include flow cytometric methods that can quantitatively detect the presence of cell surface-bound allergen-specific IgE antibodies. Assays for histamine and leukotriene C 4 released after in vitro basophil activation are now more accurate and standardized. Current analytic methods for IgE antibodies provide more quantitative and reproducible measurements of IgE than ever before, although still with less sensitivity that traditional skin testing. The current challenge is to translate the quantitative IgE antibody results into a more accurate diagnosis of allergic disease.     

Increased vascular permeability during passive peritoneal anaphylaxis in the rat. The effects of disodium cromoglycate and a nitroindanedione.

Following intraperitoneal sensitisation of rats with rat serum containing reaginic antibody, intravenous injection of blue dye and intraperitoneal challenge with antigen caused a release of histamine, slow-reacting substance of anaphylaxis (SRS-A) and dye into their peritoneal fluids. The times taken to reach peak concentrations after challenge were less than 2 min for histamine and between 5 and 10 min for SRS-A, whilst concentrations of dye were still increasing after 2 1/2 h. The amounts of histamine released by antigen were sufficient to account for about 60% of this extravasation of dye. Disodium cromoglycate (DSCG) and a nitroindanedione (BRL 10833) inhibited extravasation by inhibition of mediator release. BRL 10833, unlike DSCG, was active after oral administration, and for a given inhibition of histamine release it produced a greater effect on extravasation when given orally than when injected intraperitoneally.     

Poor association between allergen-specific serum immunoglobulin E levels, skin sensitivity and basophil degranulation: a study with recombinant birch pollen allergen Bet v 1 and an immunoglobulin E detection system measuring immunoglobulin E capable of binding to FcɛRI

BACKGROUND: Results from several studies indicate that the magnitude of immediate symptoms of type I allergy caused by allergen-induced cross-linking of high-affinity Fc epsilon receptors on effector cells (mast cells and basophils) is not always associated with allergen-specific IgE levels. OBJECTIVE: To investigate the association of results from intradermal skin testing, basophil histamine release and allergen-specific IgE, IgG1-4, IgA and IgM antibody levels in a clinical study performed in birch pollen-allergic patients (n = 18). METHODS: rBet v 1-specific IgEs were measured by quantitative CAP measurements and by using purified Fc epsilon RI-derived alpha-chain to quantify IgE capable of binding to effector cells. Bet v 1-specific IgG subclasses, IgA and IgM levels were measured by ELISA, and basophil histamine release was determined in whole blood samples. Intradermal skin testing was performed with the end-point titration method. RESULTS: Our study demonstrates on the molecular level that the concentrations of allergen-specific IgE antibodies capable of binding to Fc epsilon RI and biological sensitivities are not necessarily associated. A moderate association was found between cutaneous and basophil sensitivity. CONCLUSION: Our results highlight the quantitative discrepancies and limitations of the present diagnostic tools in allergy, even when using a single allergenic molecule. The quantity of allergen-specific serum IgE is only one component of far more complex cellular systems (i.e. basophil-based tests, skin tests) used as indirect diagnostic tests for IgE-mediated allergic sensitivity.     

Pathogenesis of esophageal rings in eosinophilic esophagitis

Eosinophilic esophagitis and eosinophilic gastroenteritis is being recognized more frequently among the adult patients. The disease is characterized by massive infiltration of the wall of gastrointestinal tract by sheets of eosinophils. The clinical features depend upon the site of involvement. They include dyspepsia, dysphagia, nausea, vomiting, chest pain, diarrhea and protein-losing enteropathy. Eosinophilic esophagitis may present as chest pain, dysphagia or dyspepsia. The characteristic endoscopic feature of eosinophilic esophagitis is the formation of fine concentric mucosal rings (corrugated esophagus). Regarding the pathogenesis of these mucosal rings our hypothesis is that mast cells in the esophageal wall in response to allergens release histamine, eosinophilic chemotactic factor and platelet activating factor, etc. which activate eosinophils to release toxic cationic proteins. Activation of acetyl choline by histamine may cause contraction of the muscle fibers in the muscularis mucosae resulting in the formation of esophageal rings. This hypothesis can be tested by demonstrating the contraction of muscle layers of muscularis mucosae with the use of high frequency endoscopic ultrasonic probe introduced via the biopsy channel of an endoscope.     

An immunological approach to the diagnosis of food sensitivity

Nineteen patients with delayed onset food sensitivity were compared with fourteen patients with immediate reactions and twenty-one non-atopic subjects in terms of clinical symptoms, foods involved and IgE mediated immunological reactions. The immediate reactors were frequently positive to all tests used: skin tests (71%), allergen induced leucocyte histamine release (71%), radioimmunodiffusion (55%) and skin window (55%). Those with the delayed onset variety were seldom positive by skin testing (13%), or skin window (0%), while 39% were positive by leucocyte histamine release and 48% demonstrated specific IgE food antibodies. Control subjects had negative responses to immunological tests for IgE antibody except for leucocyte histamine release (24%). Reasons for the differences between immediate and delayed onset food sensitivity in clinical symptoms, foods involved and immunologic parameters are discussed. A careful history in conjunction with the elimination and challenge technique remains the most useful tool at present for the delayed onset group. In vitro methods for detecting specific IgE responses may also prove to be helpful.     

A double-blind comparison of ketotifen and disodium cromoglycate in atopic adult asthmatics

The therapeutic effects of inhaled disodium cromoglycate (DSCG) and orally administered ketotifen were compared in thirty atopic asthmatics aged 15-34 years during a 22-week double-blind parallel group study. Ketotifen is a cycloheptathio-phene with experimental antihistaminic, anti-allergic and anti-anaphylactic effects equal or superior to those of DSCG. During the first 6 weeks of treatment, mean airflow meter readings increased and bronchodilator use diminished in those receiving DSCG, but no improvement was seen in those given ketotifen. In the next 10 weeks. concomitant therapy was reduced in both groups, but this reduction was greater in the group receiving DSCG. No serious adverse effects occurred. Asthma worsened after abrupt discontinuation of DSCG but not ketotifen. Although a small number of patients may have benefited from ketotifen. its effect on asthma was not comparable with that of inhaled disodium cromoglycate.