Immunomodulatory drug

 

(57) Abstract:

Usage: the invention relates to medicine and veterinary medicine, specifically to immunomodulatory drugs on the basis of hydrophobic derivatives of interferon inductors used in antiviral therapy. The objective of the invention is to expand the area of effective application, increase efficiency, reduce toxicity and to simplify the manufacture of the medicinal product. The invention: is the use of esters 10-metrocable-9-acridone as a low molecular weight interferon inducer together with a pharmaceutically acceptable carrier, the choice of means to ensure its solubility in water and microcapillary in lipid media. The technical result consists in strengthening the immune-modulating activity. 2 C.p. f-crystals, 4 PL.

The invention relates to medicine and veterinary medicine, specifically to immunomodulatory drugs on the basis of hydrophobic derivatives of interferon inductors used in antiviral therapy.

Prior art

Known used in antiviral therapy immunomodulatory drugs is acid), for example, larigan (ridalin) [1];

synthetic high-molecular inducers (polyribonucleotide acid), for example, Polivanova (poly (G), Polinesia (poly(I)) and other acids [2];

natural (vegetable) polyphenols, for example, hossipole [3];

synthetic analogues of plant polyphenols, for example, magasin [4];

synthetic small molecule inducers:

based on the 10-greenexpo acid or its sodium salt [5];

on the basis of 3,6-bis(heteroanalogs) acridine, namely 3,6-bis(2-diethylaminoethoxy) acridine [6].

Listed medicines give specific preventive and therapeutic effect. They are used dissolved in water, ointment, or other pharmaceutically acceptable carriers.

It is also known that the above tools have several disadvantages. In particular, natural and synthetic polyribonucleotide acid is highly toxic and rapidly destroyed by nucleases of the body. Natural and synthetic polyphenols typical low interferon-inducing activity and toxicity. Synthetic small molecule inducers of interferon-based 10-greenexpo acid or its sodium salt tion of the lipid layer of cell membranes and, as a result, weakens the impact of the drug to target cells of the immune system that impedes the penetration of the drug through the blood-tissue interfaces (including the blood-encephalitides) barriers. In addition, due to hydrophilicity, the drug is rapidly eliminated from the body, which requires repeated his introduction to therapeutic effect.

Synthetic small molecule inducer of interferon-based 3,6-bis(2-diethylaminoethoxy) acridine, in particular - 3,6-bis (2-ethyleneoxy)-4,5-dichloro-2-nitro acridine hydrophobic does not have the disadvantages mentioned above of the inductor based on the 10-greenexpo acid or its sodium salt. This tool is most close to the claimed its essence and mechanism of action on cells of the immune system, therefore, selected as a prototype. In more detail, the prototype described in U.S. patent N 4314061, MKI C 07 D 413/14, NCI 544-80 declared 01.08.77 published 02.02.82 (authors: K. C. Murdock, M. R. Damiani, F. E. Durr).

Along with the great advantages of the prototype, as well as other analogues, has a number of weaknesses:

a relatively narrow region of the effective action (mainly activity against herpes simplex virus-1);

high toxicity;

The present invention is to reduce these disadvantages, i.e., the expansion of the effective action, improved efficiency, reduced toxicity and simplify production.

This task is solved in that in known drug-containing low-molecular inducer of interferon and a pharmaceutically acceptable carrier, introduced significant changes, namely: as an interferon inducer selected broadcast 10-metrocable-9 - acridone.

In addition, in aqueous media, to ensure solubility in the solution can be introduced pH-stabilizing additives, for example, citrate buffer, and hydroxides of alkali and alkaline earth metals in equivalent amounts to the ethers.

In addition, when the lipid carrier, to improve the effectiveness of the solution can be microcapsulated.

The above selection is based on research conducted by the authors of the present invention, discovering new physico-chemical properties of esters 10-metrocable-9-acridone, the main of which is the high pharmacological activity and new pharmacological properties, the ability to dissolve in water in the presence of neutral or near-neutral pH values. These features and properties of esters 10-metrocable-9-acridone, unknown to the authors of the available sources of information enabled us to carry out the task.

Dosage form based on esters 10-metrocable-9-acridone includes optimal for a given pharmacological effect of the number of ester and stabilizing additives which are dissolved in pyrogen-free water (other pharmaceutically priemlimom media) or incorporated into artificial lipid structures (other pharmaceutically acceptable carriers). The inventive structure of the dosage form has allowed to obtain a set of biologically active and pharmaceutically acceptable dosage forms - parenteral, oral, inhalation, dermal, and intranasal. In other words, a new immunomodulatory agent is built on the basis of pharmaceutically acceptable complexes, esters of 10-metrocable-9-acridone with macromolecular carriers. The obtained macromolecular interferonogenna complexes have a high efficiency induction of interferons, spread spectrum antiviral activity, reduced incidence of side effects and improved pharmacist the Ira 10-metrocable-9-acridone. In particular, the use of these esters and amphiphilic lipid molecules that initially form a complex solution in acceptable organic solvents, and then, when the phased replacement of organic solvent on the pharmaceutically acceptable aqueous solution, the formation of lipid smectic mesophases (liposomes), which is an artificial spherical membrane sizes 60 to 7000 nm, consisting of one or more layers, in which the esters of 10-metrocable-9-acridone associated with the lipids of non-covalent (hydrogen and hydrophobic bonds.

The claimed medicinal product receive, for example, as follows. The mixture of lipids (0.5 and 0.9 EQ.) and of ester 10-metrocable-9-acridone (0.3 to 0.1 EQ.) directly or indirectly dissolved in an acceptable organic solvent is a chlorinated hydrocarbon, simple aliphatic esters, alcohols or other acceptable solvents. This ratio is optimal because, while the proportion of ester 0.1 - 0.3 EQ. (depending on the structure of the ether radical) stability of the formed complex is reduced and there is a crystallization of ester 10-metrocable-9-acridone. Slimming and complex. Replacement of the organic solvent in water, or by removal of solvent followed by hydration of dry matter, or by removal of the organic phase of the emulsion, the organic solvent - aqueous phase. The procedures for the replacement of the organic solvent in the aqueous phase should occur at a temperature greater than the temperature of the phase transition is the most refractory of the present in a mixture of lipids. The organic phase is determined by the solubility of the components in an acceptable solvent, and the volume of the aqueous solution is the number used for the synthesis of complex components.

Analysis of the obtained complexes using scanning and transmission electron microscopy, dynamic laser light scattering, flow cytofluorometry, liquid chromatography and high resolution column chromatography on hydrophobic columns (POLY-PREP showed that the yield homogeneous stable liposomes is at least 95% and 89% used ether stably integrated into liposomes.

The possibility of receiving the claimed medicinal products is illustrated by the following practical examples.

oC for 1 h During this time, 80 ml of distilled solvent. Cool to room temperature. To the obtained mixture is added 400 ml of pyrogen-free water and filtered. The filtrate spectrofluorometrically determine the content of ethyl ether 10 methylcarbamic-9-acridone. Add pyrogen-free water to obtain a solution of desired concentration and contribute citric acid to pH = 7,75 (approximately 1.5 g of acid). Received dosage form is filtered through a bacterial filter and poured into vials with subsequent sealing and sterilization. Get ready for parenteral use dosage form with a yield of about 98% (counting on the ethyl ester of 10-metrocable-9-acridone). Described parenteral dosage form as additives, stabilizing pH, may contain: substituted phenols, carboxylic acids, salts of weak bases and strong acids.

Example 2.

500 mg of lipids (dipalmitoylphosphatidylcholine, totaliterian in a molar ratio of 10 : 0,1) and 100 mg of lauric ester 10-metrocable-9-acridone dissolved in 500 ml of chloroform at the 22oC and placed in a thick-walled round-bottom to a thin translucent film with a greenish tint. The flask is placed in an oven equipped with a trap moisture, and finally dried in vacuum for 1 h In a flask make 10 ml of double-distilled demoninational water and after purging the flask with helium registryroot film by intensive shaking at 65oC for 1 h Produced a 10 ml thick gel with a greenish tint.

The gel is diluted to 90 ml of phosphate buffer (pH=7,4; 22oC) and treated with ultrasound at 44 kHz 30 times in 30 minute intervals. Treated with ultrasound, the suspension is forced under pressure (twice consecutively) nitrogen through polycarbonate membranes with a pore diameter of first 500, and then 100 nm. Formed opalescent suspension with a greenish tint, which is an aqueous suspension of small single-layer liposomes size 80 - 130 nm, containing 500 mg of lipids and 100 mg lauric ester 10-metrocable-9-acridone. Suspension in the presence of the stabilizer (trehalose) frozen in liquid nitrogen, and then subjected to freeze drying for 24 hours the Resulting powder is subjected to microencapsulation in acid-resistant coating. Microcapsules containing 500 mg of lipids and 100 mg lauric ester 10-metrocable-9-acridone, pressed into tablets tidalholm, the phosphatidylethanolamine, and cholesterol in a molar ratio of 1 : 0,001 : 0,5) dissolved in 400 ml of diethyl ether at a temperature of 22oC. 50 mg of ethyl ether 10 metrocable-9-acridone dissolved in 1 ml of dimethyl sulfoxide at 60oC, then cooled to 22oC and poured dropwise to 400 ml of a solution of lipids in diethyl ether with vigorous stirring. To the resulting solution add 100 ml of double-distilled water and the resulting two-phase system is treated with ultrasound to the disappearance of the phase boundary and obtain a disperse system (emulsion). The organic phase is removed with a rotary evaporator at a temperature above the transition temperature the most refractory of the present in a mixture of phospholipids (60oC) with a gradual strengthening of vacuum of 0.1 Torr (by the end of evaporation). Formed a 100 ml thick gel pale green color, which after freezing in liquid nitrogen is subjected to freeze-drying.

Formed powder pale green color with a total weight of 550 mg. Powder then mixed with 10 ml of phosphate buffer (pH = 7.0 and 22oC). Formed suspension pale green color, which is an aqueous suspension multilayer whether the illogical example 2.

Industrial applicability

In the attached tables 1 to 4 shows the results of comparative tests of the claimed medicinal product counterparts, the prototype and the control group.

Table 1 - efficacy of the claimed medicinal infection viral nature (in comparison with other drugs and prototype).

Table 2 antitumor activity of the claimed medicinal product (in comparison with other drugs and prototype).

Table 3 - efficacy of the claimed medicinal products in the development of leukemia L 1210.

Table 4 - comparative activity of different forms of the claimed medicinal product by infection with influenza virus A/Aichi/2/68(H3N2).

From table 1 it follows that the claimed product has a wider field of effective use in viral infection processes of nature, and its efficiency is several times higher than that of the prototype and even more in comparison with other shirokopolostnym drugs. Table 2 shows that the antitumor activity of the claimed drug several times more than that of the prototype and is comparable with the activity of the anti-Christ. nijaat the percentage of tumor cells in the blood in 2 times more efficient than the prototype. Table 4 shows that the index antiviral activity of water-soluble forms of the inventive means is higher than a suspension, and liposomal forms is higher than water-soluble (for example, a single injection of the drug for 1 h prior to infection with influenza A/Aichi/2/68 (H3N2).

In addition to the target of the experiments in the research process was continuously monitored the presence of adverse and toxic effects - skin irritation, mucous membranes, digestive disorders and cardiovascular systems, allergic reactions, etc., Toxicity (therapeutic index) amounted to declare funds 100 - 500, while for the prototype it was 10 to 15.

From the above examples of the preparation of a medicinal product it is clear that his production is quite simple, and the complexity of at least one and a half times lower than that of the prototype.

Thus, in our opinion, declared the drug is new, involves an inventive step (not obvious), industrially applicable and successfully solves important for mankind the task received a new high-performance, pharmaceutically acceptable vehicle for parenteral, enteral, outdoor and other applications

The letter is ractice and to industrial production, M: Ministry of health of the USSR, 1990.

2. Gordon G., M. A. Minks The imterferone renaissance: molekular aspects of induction and action. Environ. Rev., vol. 45, p. 244, 1981.

3. Hudson J. B. Antiviral compounds from plants. CRC Press, 1990.

4. Ershov, F. A. Interferon, in the book of Common and private Virology, M.: Medicine, 1982, S. 323 - 341.

5. U.S. patent N 3681360, Antiviral substitute acridones. Declared 09.04.71 published 01.06.72.

6. U.S. patent N 4314061, MKI C 07 D 413/14, NCI 544-80. Some of 3,6-bis(heteroanalogs) acridine, K. C. Murdock, R. Damiani, F. E. Durr. Declared 01.08.77 published 02.02.82.

7. Gamage I., A. Swarha, W. Gordon. Structural-activity relationship for substituted 9-oxo-9,10-dihidracridine-4-acetic-acid in J. Anti-Cancer Drug. Des., p. 403-414, 7(5), 1992.

Tareporewala J., B. Zrach et al. Synthesis and structure-activity relationship of anti-inflammatory 9,10-dihidro-oxo-2-acridinealkanoic acids and 4(2-carboxyphenyl), J. Pharm. Sci. p. 173-178, 79(2), 1990.

1. Immunomodulatory drug, containing low molecular weight hydrophobic inducer of interferon and a pharmaceutically acceptable carrier, characterized in that as an interferon inducer selected broadcast 10-metrocable-9-acridine.

2. Means under item 1, characterized in that in aqueous media in the solution introduced pH-stabilizing additives, such as citrate buffer, and hydroxides of alkali and alkaline earth metals in equival microcapsular.

 

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