RU2184544C1 - Antiallergic preparation - Google Patents

Abstract

FIELD: medicine, allergology. SUBSTANCE: invention relates to a preparation used for treatment of allergy. Invention proposes an antiallergic preparation based on quinuclidyl-3-diphenylcarbinol hydrochloride comprising additionally 1,3-dimethylxanthine taken in the definite ratio of components. Invention provides the decrease of toxicity of substances used and the combined preparation shows the preferred curative effect as compared with the known available antiallergic agents, among that preparations based on separately taken components. EFFECT: enhanced effectiveness of preparation. 3 tbl, 7 ex

Description

 The invention relates to medicine and can be used as a medicine for the treatment of allergic diseases.

 H1 antihistamines are most widely and long-term used in antiallergic therapy (1): diphenhydramine, suprastin, pyrilamine, pipolfen, ketotifen and others. Having a sufficiently high therapeutic efficacy, the above-mentioned histamine H1 receptor antagonists cause a number of undesirable side effects that often limit their use. In addition, drugs of this class are characterized by a decrease in therapeutic efficacy (tachyphylaxis) with prolonged use.

 It is known in medical practice to use drugs (2) that inhibit the allergic activation of cells and the secretion of allergy mediators from them by affecting intracellular processes of regulation of cellular functions by changing the level of accumulation of intracellular messengers (transmitters). An example of such drugs are xanthine derivatives, which have antiphosphodiesterase action, cause an increase in the intracellular content of cyclic 3,5-adenosine monophosphate (cAMP), which leads to inhibition of allergy mediators. However, these drugs do not have a noticeable effect on the activation of histamine, which plays a crucial role in the development of all known external manifestations of the allergic process, and therefore is not effective enough in the treatment of a number of allergic diseases. In addition, preparations based on xanthine derivatives have a fairly high toxicity, which also does not contribute to their mass use for the treatment of allergies.

 Since an allergic reaction is a complex cellular organization process, including many mechanisms for activating the secretion of various allergy mediators, providing the possibility of influencing several factors of an allergic reaction is one of the ways to improve the treatment of allergic diseases, including increasing the effectiveness of pharmacological treatment by creating multifunctional drugs .

 A directed search in this direction and the creation of fundamentally new chemical compounds with a multifunctional effect (3) is most desirable, but it is a long, complex and expensive process, and therefore the solution of this problem based on combinations of economically acceptable, affordable and highly effective drugs is most feasible in the present conditions .

Of the known H1 antihistamines, by their technical nature and by the effect exerted, the closest to the present invention is phencarol, an antiallergic drug (4), including quinuclidyl-3-diphenylcarbinol hydrochloride-C ₂₀ H ₂₃ NO HC1 (CDFA). The well-known drug has a pronounced antihistamine activity due to the high selectivity of the blockade of histamine H1 receptors and does not have undesirable side effects (sedative, hypnotic effects, effects on the cardiovascular system, gastrointestinal tract, vision, urinary system, etc.) specific to the above H1 antihistamine drugs.

 However, the therapeutic effectiveness of the known drug, like any antihistamine, is limited by its ability to suppress only the action of histamine. In addition, HDPC affects not only an allergic reaction, but also causes an antiproliferative effect, which significantly reduces its antiallergic effect.

 Thus, the technical result obtained from the implementation of the described invention consists in creating, on the basis of available and low cost source components, a new, multifunctional, highly effective antiallergic drug.

The specified technical result is achieved in that the antiallergic drug, including quinuclidyl-3-diphenylcarbinol hydrochloride, additionally contains 1,3-dimethylxanthine in a mass ratio of components:
quinuclidyl-3-diphenylcarbinol hydrochloride: 1,3-dimethylxanthine 1 ÷ 2: 4.

1,3-dimethylxanthine - C ₇ H ₈ N ₄ O ₂ (DMK) belongs to the group of methylxanthines and, in its own use, theophylline, has a vasodilating, bronchodilator and antiallergic effect, which is explained by the variety of its pharmacological activities, manifested on cellular and molecular levels. So, in addition to antiphosphodiesterase activity, DMC interferes with the purine energy system, blocking the action of adenosine, and enhances apoptosis (including cells involved in allergic inflammation). As a result, the anti-allergic effect of DMC can be associated with a complex of effects: with inhibition of histamine secretion from mast cells and basophils, with inhibition of respiratory explosion in eosinophils, with some inhibition of prostaglandin synthesis and degranulation of eosymnophils, inhibition of proliferation of T-lymphocytes and interleukin 2 production.

 The combined preparation in accordance with the present invention is superior in antianaphylactic activity, bronchodilator and antiexudative action to each individual substance, and also has a lower toxicity. The described preparation has advantages in the ability to inhibit the secretion of mediators from allergy target cells - basophils. In addition, for the first time, the ability of DMK to significantly reduce the antiproliferative effect of HDPC was found, which further indicates the advantages of the combined drug.

 The effect of the combined drug is due to the effect on the allergic process both by blocking the histamine H1 receptors and by inhibiting the function of target cells and thereby preventing the secretion of various allergy mediators from them.

 The antiallergic drug (theophene) according to the invention can be prepared by mixing powdered HDPC and DMK in a ratio of 1 ÷ 2: 4 with the addition, depending on the type of dosage form, of auxiliary substances that are selected from the group of carriers, preservatives and / or flavorings. For example, for the preparation of the drug in the form of tablets, you can use excipients such as starch, various sugars, calcium phosphates.

 The tablets receive a white or white with a yellowish tint.

 The drug is slightly soluble in water and can be dissolved in solutions of acids, for example, hydrochloric acid, as well as alkalis.

An example of the use of the described drug are tablets in the composition, g:
1,3-Dimethylxanthine - 0.1
Quinuclidyl-3-diphenylcarbinol hydrochloride - 0.025
Corn starch - 0,111
Milk sugar - 0.0621
Calcium Stearate - 0.003
The biological activity of the described drug was studied in comparison with the drugs fenkarol and theophylline.

 The following examples show the methods and results of testing the properties of the described anti-allergic drug.

 Example 1. The study of the antiallergic effect of the drug.

 A. In vivo model of active anaphylaxis.

 The study was carried out on actively sensitized rats of both sexes weighing 180 ÷ 200 grams with oral administration of the drug 2 hours before the introduction of a resolving dose of antigen. Evaluation of the effectiveness of anaphylactic reactions was carried out by plethysmometric registration of increment of the foot volume of the hind paw of rats.

 Antigen-induced edema of the paw of rats was chosen as the main model, as the most fully reflecting all stages of the development of an allergic reaction in the whole organism.

It was found that the average effective antiexudative dose of the combined preparation according to the invention, i.e. the dose that suppresses paw edema in rats is 50% (ED ₅₀ ), significantly lower than each of its components when used separately, which indicates its predominant anti-allergic activity.

 The research results are presented in table 1 (1).

 B. In vivo model of passive anaphylaxis.

 The studies were carried out on intradermally sensitized male rats weighing 300–350 g by oral administration of “theophene” at a rate of 30, 60, and 90 mcg / kg 1–1.5 hours before the resolving injection of the allergen. Evaluation of the effectiveness of the drug was carried out by determining the severity of passive cutaneous anaphylaxis (PKA) when comparing the average values of the logarithms of reciprocal titers by the intensity of the PKA according to the Fox criterion.

 The results are shown in table 2, confirm a significant decrease in the degree of intensity of PKA with the introduction of the drug "Theofen" compared with the control.

 Example 2. The study of bronchodilator activity.

 Studies were conducted on anesthetized guinea pigs with intravenous histamine (10 mcg / kg). An increase in the tone of the bronchial muscles was recorded according to the method of Konzett, Rossler in the modification of M.E. Fireplaces. The studied drugs were administered 3 minutes before the injection of histamine. The bronchodilator activity of the drugs was judged by their ability to reduce the amount of histamine-induced bronchospasm.

 The results obtained indicate the potentiation of the bronchodilator action of the described drug.

 The research results are presented in table 1 (II).

 Example 3. The study of antiexudative activity.

 The studies were conducted according to the method of evaluating histamine-induced ophthalmic reaction in outbred guinea pigs of both sexes weighing 250–350 g. Conjunctival edema was performed by instillation of two drops of a 2% solution of histamine dihydrochloride. The studied drugs were administered orally in the form of an aqueous suspension 1 hour before the installation of histamine.

 The effect of the drug was judged by the state of hyperemia of the mucous membrane of the eyelid.

 Listed in table 1 (III), the research results show the predominant antiexudative effect of the described combination drug "Theofen."

 Example 4

 Assessment of the effect of the drug on the level of intracellular cAMP content.

A comparative study of the effect of the described drug on adenosine-stimulated increase in the level of cAMP in lymphocytes was carried out. The final concentration of adenosine in the samples was 0.1 μM, which corresponds to the physiological concentration of purine in the blood. The content of cAMP in lymphocytes was assessed using the radioligand method. The cell concentration in suspension was 3 million / ml of 199 medium containing 10 mM HEPES. Cells were incubated at a temperature of 37 ^° C. The test preparations were added in a volume not exceeding 50 μl. At the end, the cells were destroyed by heating at a temperature of 90 ^{° C.} for 2 minutes, centrifuged at 3000 g for 5 minutes, and 200 μl of the supernatant were taken for analysis of cAMP content. The concentration of the cyclic nucleotide was expressed in pmol / 10 ⁷ cells ± a significant error of the mean at a significance level of p <0.05 according to the results of 7 ÷ 8 independent experiments.

 The research results are shown in table 1 (IV).

 Studies of the action of the described drug showed a significant increase in the increase in the level of intracellular cAMP in lymphocytes caused by adenosine compared with individual components.

 Example 5. The study of the effect of the drug on the effector phase of allergies.

 Studies of the action of the drug were carried out according to the reaction of histamine release from basophils of human peripheral blood in vitro.

 We used peripheral blood samples from healthy donors and patients aged 18 to 47 years with increased sensitivity to pollen from grasses and trees.

To determine the effect of the drug on the allergen-induced histamine release reaction, the cell suspension was incubated without or in the presence of test agents for 40 min at a temperature of 37 ^° C. Then, the allergen was added to the samples and the incubation was extended for another 40 min. The histamine content was determined by microspectrofluorometric method in sedimentary portions of cells without preliminary extraction of histamine. In order to exclude the influence of individual differences in the degree of allergic release of histamine from basophils, the results of the experiments were expressed as a percentage of the maximum release of histamine in this test on the basophils of this patient.

 Studies have shown that HDPC at a concentration of 10 μM and DMK at a concentration of up to 100 μM did not statistically significantly affect the secretion of histamine caused by an allergen at a concentration of 0, l PNU / ml. The exposure of the described combination drug caused inhibition of the allergic release of histamine at about the same allergen concentration, which was approximately twice that in the control group, and to achieve a similar effect when used separately, a five-fold higher concentration of HDPC was required.

 The test results are presented in table 3, which illustrates the pronounced potentiation of the inhibition of submaximal allergic histamine release in vitro tests.

 Example 6. The study of the effect of the drug on proliferation.

Studies were performed by evaluating the proliferative activity of peripheral blood lymphocytes in patients with atopy. For this purpose, mononuclear cells were isolated from the peripheral blood of patients with atopy by centrifugation on a single-stage density gradient of ficollvergrafin (1080 g / cm). As a coagulant, a 2.7% solution of sodium ethylenediaminotetraacetate was used in a ratio of 1 ml per 10 ml of blood. Mononuclear cells isolated and washed with buffer solution were cultured in RPMI 1640 medium containing 10% serum of cow embryo, 5 mM HEPES, 20 mM glutamine and 100 U / ml gentamicin. Phytohemagglutinin (PHA, Sigma) was added to the cells at a final concentration of 1.5 x 10 μg / ml and test preparations. The cultivation was carried out in an atmosphere with high humidity and CO ₂ content in air for 2 hours. The result was evaluated by the inclusion of H ³ thymidine, introduced 4 hours before the end of the incubation.

As a result of the study of the influence of the studied drugs on the mitogen-induced proliferative reaction, it was found that:
- the level of proliferation of mononuclear cells under the action of HDPC at concentrations of 10 ^-3 M, 10 ^- M and DMK at concentrations of 10 ^-4 M was, respectively, 2680 imp / min, 4750 imp / min and 4950 imp / min, which indicates almost complete suppression of the proliferative response of immunocompetent cells,
- at concentrations of 10 ^-5 M, the compounds of HDPC and DMK did not significantly affect proliferation,
- the level of proliferation under the action of the combined drug "Theofen" amounted to 29570 + 3200 cpm / min, which indicates a weakening of the antiproliferative effect of independently used HDPC and DMK.

 Example 7. Acute toxicity test.

 Acute toxicity of the combined preparation was evaluated in 4 groups of outbred white mice weighing 18-20 g. The preparation was administered Per os in 0.3 ml of aqueous suspension based on the preparation 100, 200, 400, 800 mg / kg. For each dose of the drug there were 5 animals. Observation of animals and registration of their death was carried out for 14 days. The death of animals occurred within only the first two days after administration of the drug. No features were observed in the state of the surviving animals. The reproducibility of the obtained results was tested on a repeat experiment similar to a 10-day interval test performed on outbred white male mice with doses of theophene - 200, 300, 600, and 900 mg / kg.

LD _{50 was} calculated according to the Kerber formula.

 The results obtained are presented in table 1 (IV).

 It was found that the toxicity of the combined drug is reduced compared to each individual substance and amounts to 455 mg / kg, which allows it to be classified as moderately or slightly toxic drugs. Moreover, in combination with HDPC, a sharp decrease in the toxicity of DMC is observed.

 Thus, the described combination drug, having a multifunctional effect, surpasses the main indicators that determine the antiallergic properties of the drug, its components when used independently and is less toxic, which makes it possible to recommend it for treatment with greater effectiveness for a wide range of allergic diseases.

Sources of information
1. Zarudy F.S., Histamine and antihistamines, Ufa, 244s.

 2. EP, application 344586, cl. MKI A 61 K 31/71, op. 1989.

 3. USSR author's certificate 1100875, MKI C 07 D 473/08, op. 1986 year

 4. Mashkovsky et al., "Fenkarol and its use in the treatment of allergic diseases", g. Clinical Medicine, 1978, 11, p. 22-28.

Claims (1)

 Antiallergic drug, including quinuclidyl-3-diphenylcarbinol hydrochloride, characterized in that it additionally contains 1,3-dimethylxanthine in a mass ratio of components - quinuclidyl-3-diphenylcarbinol hydrochloride: 1,3-dimethylxanthine = 1-2: 4.

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