WO2003043629A1

WO2003043629A1 - Medicine based on diethylaminoethanol derivatives and its use for the prevention and treatment of inflammatory and degenerative diseases

Info

Publication number: WO2003043629A1
Application number: PCT/RO2001/000022
Authority: WO
Inventors: Alexandru Vrabiescu
Original assignee: Alexandru Vrabiescu
Priority date: 2001-11-22
Filing date: 2001-11-22
Publication date: 2003-05-30
Also published as: AU2002224253A1; EP1490046A1

Abstract

The derivatives of diethylaminoethanol are esters of the phthalic acid or of the aminophthalic acid, according to the formula (I) where X is H or a -NH2 group, or a pharmaceutically acceptable salt thereof. The above-mentioned derivatives produce antiinflammatory, immunomodulatory and antioxidant effects that are stronger than those of diethylaminoethanol, also having a much lower toxicity. The medicine for the prevention or treatment of inflammatory or degenerative diseases contains, as active ingredients, esters of diethylaminoethanol with phthalic acid or aminophtalic acid. The use of the medicine of the invention consists in its administering in the prevention or treatment of inflammatory and degenerative diseases.

Description

MEDICINE BASED ON DIETHYLAMINOETHANOL DERIVATIVES AND ITS USE FOR THE PREVENTION AND TREATMENT OF INFLAMMATORY

AND DEGENERATIVE DISEASES

This invention refers to a medicine based on diethylaminoethanol derivatives and its use for the prevention and treatment of inflammatory and degenerative diseases.

Human therapy makes use of numerous substances in order to combat and prevent inflammatoiy and degenerative diseases.

Thus, antibiotics are widely used in the treatment of inflammatory diseases; they have saved many lives or have greatly improved the condition of the patients. However, their extensive and sometimes unjustified use resulted in the appearance of germ strains that resist treatment with the known antibiotics. Consequently, it has become necessary to increase the dosage of the administered antibiotics and that increased the risk of toxic effects to occur.

At the same time, there appeared diseases produced by infectious agents that resist to antibiotics and frequently have a serious evolution.

Under these conditions, there is and increased interest in finding new antiinflammatory substances, more efficient than the known ones and that would not generate side effects or would have only minor side effects.

At present non-steroid anti-inflammatory substances such as those in the classes of indolic. arylacetie, propionic, arylpropionic and oxicam derivatives are widely used.

These substances basically act by inhibiting the biosynthesis and release of prosta^andin in the peripheral tissues. This inhibitory action is, in fact, the cause of most of the side effects that may to appear in the course of treatment, such as: Digestive disorders, e.g. nausea, diarrhoea, epigastric pains, gastro-intestinal ulceration, haemorrhage, perforations; cardio-vascular disorders, e.g. hypertension bouts, oedema; haematological disorders, e.g. leucopoenia, thrombocytopoenia, agranulocytosis, medullar aplasia; neurological disorders, e.g. asthenia, insomnia, headache; renal disorders, e.g. renal failure, nephrotic syndrome; sight and hearing disorders.

Several substances having immunomodulatory properties are also used in the treatment of inflammatory diseases. Examples of such substances are imidazothiazol, molgramostim, filgrastim, A-interferon, azathioprinum, 4-iodantipyrin, butadion, sodium salicylate, dinitrochlorobenzen, cyclosporins.

The major disadvantage, involved in the employment of those substances for more than 30 years, has been that their utilization for therapeutic purposes did not go beyond a limited number of cases. The reasons were the mediocre outcome from their use, the restrictions imposed by their toxicity, their side effects and the methodological or economic difficulties involved in employing them.

None of these substances has proved able to stimulate the body's defences to such an extent as to make them usable in reducing the incidence or seriousness of inflammatory diseases in large populations or communities, as in the case of vaccines.

This disadvantage is mainly the result of their mechanism of action. Thus, some immunomodulators induce immunosuppression in the bone marrow, thus decreasing the body's defence ability, while others specifically interfere with poorly performing cellular or humoural immune mechanisms. The only products that have a higher efficiency are those based on interferon, but their use is much limited because of their toxic effect and of their very high cost.

The side and toxic effects they induce represent another disadvantage of many of the immunomodulatory substances known to date. Thus, imidazothiazol may induce nausea, vomiting, diarrhoea, stomatitis, taste and smell alterations, rashes, increase of body temperature, alopecia, leucopoenia, arthralgia and myalgia. The most common haematological side effect is agranulocytosis, followed by leucopoenia and thrσmbocytopenia, all of them likely to lead to serious complications.

Special attention was given to the use of cytokines in the treatment of several inflammatory diseases. Such methods include the oral or parenteral administration of large amounts of cytokines. This is not only very costly but it can also induce general metabolic disturbances. External application is inefficient because of decomposition or poor absorption.

The only products having a greater efficiency are those based on interferon. However, their use is much restricted because of their high cost and also because of their toxic effects.

Administration of A-interferon may induce, in most of the patients, flew-like symptoms, fatigue, fever, shivering, anorexia, myalgia, headaches, arthralgia, perspiration; in two thirds of the patients anorexia may appear and half of them complain of nausea. There have also been reports of rashes, drying of the skin, alopecia, rhinorea, epistaxis.

Antioxidant agents have also been used in the treatment of many inflammatory and degenerative diseases frequently accompanied by a significant increase of oxygen free radicals.

A series of vitamins such as vitamin C, vitamin E, beta-carotene, vegetal or mineral products, such as selenium and zinc, have been used as antioxidants in the treatment of diseases with a severe evolution, e.g. AIDS, therapy-resistant chronic inflammatory diseases, cancer, neurological and cardio-vascular degenerative diseases.

The low efficacy of those therapies explains the constant search for new antioxidants or substances able to stimulate the antioxidant mechanisms of the body. Procaine and pharmaceutical products based on procaine are now being more extensively used in the treatment of inflammatory and degenerative diseases, particularly in the elderly.

Also, much research has been performed in order to replicate the therapeutic effects of procaine in experimental models and understand its mechanisms of action. During experimental research, it was found that effects were much better with doses by unit of body weight that were ten times greater than those regularly used in humans.

Therefore, an increase of therapeutical dosage in humans could produce better clinical effects, easily identified by laboratory tests, and in a shorter time.

The disadvantage lies in the fact that, in humans, the procaine dose administered parenterally cannot be increased without mcurriπg the risk of side effects such as dizziness and nausea, that require in most of the cases the discontinuation of treatment.

Another disadvantage of the clinical use of procaine consists in seπsitisatioπ to procaine accompanied by adverse reactions such as headaches, trembling, cramps in the exfremities, tachycardia or bradycardia, motor excitation phenomena.

Recently published experimental research brings new data on the biological properties of diethylaminoethanol (DEAE), one of the products of procaine hydrolysis, considered to be the most active factor of this substance.

Thus, good effects have been obtained in the treatment with DEAE in rats with arthritis induced with Freund adjuvant (VRABIESCU AL and coll., Rom. J. Physiol., 1998, vol. 35, 1-2, p. 99-110).

Other research refers to changes produced in the immune system (BORDEA M. and coll., Rom. J. Physiol., 1998, vol. 35, 1-2, p. 111-126; VRABIESCU AL. and coll., Rom. Arch. Ivficrobiol. Immunol., 1998, vol. 57, 2, p.l 11-124).

Thus, in rabbits, one found an increase in the circulating lymphocytes count and also histoptahological changes in the sense of a stimulation of lymphoid structures.

Other research pointed out that DEAE treatment can to a certain extent influence the release of oxygen free radicals by the polymorphonuclear cells. (DOLGANIUC A and coll., Rom. Arch. Microbiol. Immunol., 1998, vol. 51, 1, p. 23-32).

It is worth mentioning that the DEAE doses administered during the above-mentioned studies have been larger than the amount of DEAE contained in a dose of procaine used in therapy.

That was possible because of the much lower toxicity of DEAE (DL₅₀=1300) compared to procaine (DL„o⁼300).

The information presented above points to the special interest shown for the identification of new therapeutic means. This interest is currently enhanced by the unprecedented increase in the frequency and seriousness of inflammatory and degenerative diseases and by the appearance of widely spread diseases that are difficult to treat and that result in much suffering, the reduction of the work capacity and impairment of social life, disabilities and a shortening of the duration of life.

To that, one should add the increased health care expenses and the decrease in the living standards. This also results in an increase in medicine consumption, in the number of hospitalisation days and the number of medical and surgical procedures that need to be performed. There is a negative impact on the health insurance funds that cannot be used for the development of research and improvement of health care services.

Under such conditions, the development of medicines with a higher efficiency in combating inflammatory and degenerative diseases is of particular interest.

The problem solved by this invention is that of developing a medicine that could significantly enhance the antiinflammatory, immunomodulatory and antioxidant properties of dielJ ylaminoethanol described above, so as to produce safe therapeutic effects unaccompanied by side effects, or with a nώiimum of side effects.

The invention consists in a medicine whose active principle is an ester of diethylaminoethanol with the phthalic or aminophthalic acid, with the formula (1):

where X is H or a -NH2 group or a salt thereof that is pharmaceutically-acceptable and that is efficient in the prevention or treatment of inflammatory and degenerative diseases, in immune failure and in pathological conditions with an increased production of autoantibodies, immune complexes and oxygen free radicals.

The invention also refers to the use of the medicine of the invention: - in the form of capsules, pills, dragees, tablets, ampoules for the stimulation of the immune system and the treatment of inflammatory diseases and of chronic degenerative diseases; - in the form of capsules, pills, dragees, tablets, ampoules, geL lotion, cream, ointment for the treatment of rheumatoid arthritis or other rheumatic processes; - in the form of lotion, gel, cream, ointment, powder, suppositories for the treatment of skin and mucosa inflammation; - in the form of capsules, dragees, tablets, syrup, ampoules, spray, aerosol solution for the treatment of airways inflammation.

The doses used are of 5...150 mgkg bw/day, preferably 30 mgkg bw/day given orally for intervals of 15-30 days, followed by a break of 15-45 days, for prevention purposes. For curative purposes the doses are of 10...100 mg kg bw/day, preferably 30 mg kg bw/day given parenterally, or 15...20O mg/kg bw/day, preferably 60 mg/kg bw/day, given orally and/or in a concentration of 1...80%, preferably 10 g%, in local applications, for 3-20 days.

This invention consisting in the development of a medicine of the invention, cont_tining as an active ingredient an ester of diethylaminoethanol with the phthalic or aminophthalic acid, as per formula (1) and a treatment method of the invention, provides the following advantages:

• The antiinflainmatory, irnmunomodulatory and antioxidant properties of diethylaminoethanol are much enhanced, which makes the medicine of the invention particularly eflfective for therapeutic purposes.

• For the first time, a medicine has been developed that can induce (reversible) structural and functional changes in the immune system by means of mechanisms that converge and create fevourable conditions for a highly effective impact on inflamrnatory and degenerative diseases.

• Unlike non-steroid antϋnflamrnatory drugs known to date, the medicine of the invention, acts by: blocking the release of super oxide anions at the site of inflammation, tensifying the activity of antiinfl-immatory cytokines and activating the T lymphocytes. Owing to those mechanisms, the medicine, as per the invention, can stop and suppress the inflammatory process and provide an enhanced therapeutic efficacy.

• Because of its pham codynamic properties, the medicine of the invention has the ability to have a beneficial influence on diseases that are difficult or impossible to treat with the existing medicines. Among them one could mention pathological conditions involving an increase production of super oxide anions, autoantibodies and immune complexes.

» The medicine of the invention can be used in preventing inflammatory and degenerative diseases because of its ability to stimulate lymphoid structures to generate and maintain a high level of active immune elements for a long time.

• It does not produce the secondary effects of the type described for nonsteroid antiiπ_ mmatory drugs, as it acts through other mechanisms of action.

• Its toxicity is much lower than that of diethylaminoethanol, which makes it possible to administer it in much larger doses. Thus, DL₅0 is 9000 mg/kg bw in the medicine of the invention, while in the case of diethylaminoethanol chlorhydrate it is 1300 mg/kg bw. Also, the medicine of the invention, has no cytotoxic effects, is not antigenic and does not generate addiction, in therapeutic doses.

• It can be efficient in preventing or diiriinishing the secondary effects on the immune system of treatments with extremely aggressive chemical or physical agents such as cytostatics, cortisone, gamma radiations etc., used in many serious diseases for which there are no other therapeutic means. • The medicine of the invention is easy to administer as it is specially formulated for the oral and transcutaneous pathway, and it is only in specific cases that the parenteral pathway is used.

• The medicine of the invention will be widely accessible, as its preparation and conditioning do not involve high costs.

It is possible that other therapeutic effects would be added to the antiinflarninatory, immunomodulatory and antioxidant effects described above. Such effects may result from the pharmacodyiiamic actions described in literature in the case of other derivatives of the phthalic acid, i.e.: the hvpolipemic action (LAKE B.G., GRAY T.J., GANGOLLI S.D., Environ Health Perspect 1986 Aug; 67:283-90; FflNTON R.H., MITCHELL F.E., MANN A, CHSCOE D., PRICE S.C.. NUNN A, GRASSO P., BRIDGE J.W., Environ Health Perspect 1986 Dec; 70:195-210; OISHI S., HE AGA K, Toxicol Lett 1982 Nov; 14(l-2):79-84), - the cholesterolgenesis inhibiting action (BELL F.P., Environ Health Perspect 1982 Nov; 45:41-50; NAIR N., KURUP C.K., Biochem Pharmacol 1986 Oct; 15; 35(20):3441-7), - the action of protecting the arterial wall against lipidic iiifiltration (NUMANO F, The 11^th International Congress of Gerontology. Abstracts for plenary sessions & Symposia, August 20-25, 1978, p.175, Tokyo, Japan).

The medicine of the invention is formulated as capsules, pills, tablets, dragees, injectable solutions, gel, cream, ointment, lotion, syrup, powder, suppositories, solutions and the like and is administered for:

• Prevention and treatment of inflammatory or chronic degenerative diseases, of immune failure and pathological conditions with an increased production of free oxygen radicals, auto antibodies and immune complexes, when the medicine is administered in the form of capsules, pills, dragees, syrup, suppositories, injectable solutions and the like;

• Treatment of rheumatoid arthritis or other rheumatic processes, such as arthritis, spondilarthritis, myositis, tendinitis, lombosciatica, when the medicine is administered in the form of capsules, dragees, tablets, syrup, injectable solutions, jell, solution, cream, ointment and the like;

• Treatment of inflammatory diseases of the airways, of skin and mucosa inflammation and of inflamed venous stasis, when treatment is administered, depending on the diseases, in the form of solutions, lotion, syrup, spray, gel, cream, powder, suppository, capsules, dragees, injectable solutions and the like.

Pharmaceutical forms for oral administration include solid preparations such as tablets, granules, pills, dragees, powders, hard capsules and the like.

Liquid preparations include syrups, elixirs, soft capsules and the like, Syrups can be prepared by dissolving the active ingredient or its pharmaceutically-acceptable salt in an aqueous solution comprising earboxymethyl cellulose, D-sorbitol, sugar and the like, together with a preservative such as propyl paral ydroxybenzoate.

Elixirs can be prepared by mixing the active ingredient or its phaπnaceutically-acceptable salt with glycerine, propilenglycol, orange oil, lemon oil, coriander oil, natiium borate, talcum and the like.

Soft capsules can be prepared by dissolving or suspending of the active ingredient or its pharmaceutically-acceptable salt in a lipid-based solution, such as vegetable oils, oily emulsions, glycols and the like and filling soft capsules with resulting solution.

Injectable preparations can be prepared by dissolving the active ingredient or its phaπnaceutically-acceptable salt in a physiological saline, followed by filling ampoules or vials under genn-free conditions.

Locally or topical applications can include semi-solid preparations such as ointments, gels, creams and the like or in liquid preparations such as lotions, liquids for cataplasm, sprays, liniments, and the like.

The dose of the medicine of the invention may vary, depending on the nature and extend of the disease, the age and the body weight of the patient, the administration route.

In general, the unit dose for oral administration or by injection may be from 100-lOOOmg and can be aαministered 1-3/day.

For local administration, transdemiically, the unit dose is from 5 to 500 mg and can be applied 1-6/day.

However, the invention is not limited to those embodiments, and various changes and modifications may be effected therein by one skilled in the art not departing from the scope or spirit of the invention as defined in the appended claims.

The derivatives of the diethylaminoethanol of the present invention can also be used as cell culture reagents for scientific studies, experiments and therapeutical purposes, and also as active ingredients in ordinary medicines and cosmetics.

Herein below, two examples and seven biological tests are presented. The purpose is to illustrate the invention and not to limit it.

Example 1:

Obtaining the diester [2-(diethyIarnino)-ethyl] of the dichlorhydrate phthalic acid.M=153.65.

Raw materials: diethylaminoethanol = 21 ml (157.6 mmol); phthaloil chloride = 11.2 ml (77.8 mmol); toluen = 90 ml; acetone ~ 700 ml; chloroform; active charcoal. Phthaloil chloride and toluen are poured into the reaction flask. The flask is then heated to 50-80° C and the drops of dietliylamiπoethaπol are added at an extremely slow rate under mechanical stirring.

At the end, the product becomes separated in the form of a brown-reddish, viscous oil. Toluen is decanted and the product is redissolved in chloroform (40 ml), under heat. 300 ml of acetone is added.

It is left to precipitate slowly, at an average temperature (~15°C), possibly with insemination or stirred with a wand. 50 ml of acetone is added and then it is left to precipitate for 2 days.

It is the filtered and washed with acetone. It is redissolved in chloroform with acetone (5:1) under heat, treated with charcoal and filtered under heat.

300 ml of acetone is added and it is left to precipitate. It is filtered and dried (under vacuum) in an exicator on silicagel. The filtered solutions are left to precipitate and are filtered again.

One gets 14 g of substance, in the form of white crystals, soluble in water, not soluble in alcohol, ether, chloroform. The output is 41%.

Example 2:

Obtaining the ester 4-amino 1.2-diethylaminoethylphthalat chlorhydrate. M.W. = 490

Raw materials: 181 g 4-nitrophthalic anhydride, 450 g diethylaminoethanol, anhydric ethylic alcohol, benzene, concentrate sulphuric acid, solution of 0.2 moϊ of sodium etoxide, Ni-Raney, hydrazine, chloroform, anhydric acetic acid, gaseous hydrochloric acid, bleaching charcoal.

A mixture of 181 g 4-nitrophthalic anliydride, 368 g anhydric ethylic alcohol, 400 ml benzene and 10 g concentrate sulphuric acid are boiled with an ASTM water separator until no more water is separated - about 8 hours.

A sample that has been washed with water and dried, therefore a solution of ester in benzene, is IR-tested; it should no longer have any COOH groups; if it still has any, it is boiled for another 4 hours.

The solution obtained as above is concentrated to one quarter by distillation. One adds a solution of 0.2 mol of sodium etoxide (prepared by dissolving 4.6 g of sodium in 90 ml anhydric ethylic alcohol) and 450 g diethylaminoethanol and then one performs distillation in a 30 cm distillation column: the existing ethanol is quickly distilled, while the ethanol formed by transesterification is distilled slowly, in a couple of hours. The reaction is completed when nothing more is distilled, i.e. after 4-5 hours.

The product is NMR-tested: it should not contain the ethyl group.

The alcoxide is neutralized with anhydric acetic acid and the diethylaminoethanol in excess is distilled under vacuum, up to 90°/5mm. In IR, the alcohol band is no longer present. The residue is dissolved in 1 litre of anhydrous ethylic alcohol; Ni-Raney is added (about 5 g_ previously washed with alcohol by decantation). It is then heated to boiling point under stirring and 3 molls of hydrazine are added by pipetting (under the form of hydrate solution 85-95%); an intensive release of nitrogen occurs.

Stirring continues for another hour while keeping the solution warm, then the solution is filtered and alcohol is evaporated on water bath, under vacuum (80°/5mm). The residue is IR-tested (the NH2 group bands appear) and NMR-tested.

Formation of the hydrochlorate: the oily, viscous residue is dissolved in 750-1000 ml of chloroform, the solution is treated with 5-10 g bleaching charcoal, it is stirred for an hour at 50° and then filtered.

In this solution, dry, gaseous hydrochloric acid is barbotated (possibly with cooling) to the point of saturation. Hydro chlorate crystals are formed. They are filtered and washed with anhydrous ether.

If necessary, it is re-crystallized from chloroform/acetone.

At the end, the substance is in the form of white, water-soluble crystals.

The derivatives obtained according to 1 and 2 examples, mentioned above, are conditioned in the form of: - capsules, pills, tablets, dragees and the like, containing 50...500 mg of active ingredient, preferably 300 mg; - ampoules, containing 1...20 g% of active ingredient, preferably 2 g%; - solutions for ionisation and ultrasounds, containing 1...50 g% of active ingredient, preferably 20 g%; - aerosol solution, containing 0.5...10% of active ingredient, preferably 1 g%; - jell, cream, ointment, containing 1...50 g% of active ingredient, preferably 20 g%; - lotion, spray, syrup, containing 1...20 g% of active ingredient, preferably 5 g%; - drops, containing 1...5 g% of active ingredient, preferably 2 g%; - suppositories, containing 1...50 g% of active ingredient, preferably 10 g%; - powders, mixed with talcum in a 1/1 proportion; - nasal and oral solution containing 1... 10 g% of active ingredient, preferably 2 g%.

According the use of the medicine of the invention, it is administered in the form of: - capsules, pills, dragees, ampoules and the like, for the stimulation of the immune system, and for the treatment of chronic degenerative diseases; - capsules, dragees, tablets, ampoules, jell, solution, cream, ointment and the like for the treatment of rheumatoid arthritis or of other rheumatic processes such as arthritis, spon lilarthritis, myositis, lombosciatica ; - lotion, jell, solutions, syrup, powder, suppositories and the like for the treatment of skin and mucosa inflammation; - capsules, dragees, tablets, ampoules, syrup, aerosol solution and the like for the treatment of airways inflammation. Biological Tests:

Test 1:

The immunomodulatory action demonstrated by the influence on the circulating leukocyte count. The study as performed in 20 Schinchilϊa, male rabbits, weighing 1800-2000 g. One monitored the increase of the circulating lymphocyte count after administering the medicine of the invention, example 1, compared to the increase resulting from the administration of diethylaminoethanol under the same conditions.

The animals were divided into two equal groups: 1) treated with the medicine of the invention, example 1, and 2) treated with diehylaminoethanol. Treatment consisted in the daily i.m. injecting of 15mg/kg bw of the respective substance for 5 consecutive weeks.

The effect of treatment was assessed by deteπnining the leukocyte count and comparing it with the lymphocyte count in peripheral blood after 3 and, respectively, 5 weeks of treatment, with the count before initiation of treatment, hi this manner, every rabbit served as its own control.

Tables 1 and 2 show the results obtained after calculating the group average after 3 or 5 weeks of treatment.

Table 1 : Increase in the circulating lymphocyte count in rabbits treated for 3 weeks with the medicine of the invention compared to rabbits treated with diethylaminoethanol.

*The medicine of the invention, example 1

Table 2: Increase in the circulating lymphocyte count in rabbits treated for 5 weeks with the medicine of the invention compared to rabbits treated with diethylaminoethanol.

"The medicine of the invention, example 1. One finds that in the group treated with the medicine of the invention, example 1, the lymphocyte count increased by 45.1% (pθ.0006) after the first 3 weeks of treatment, unlike in the rabbits in the ctøethylaminoethanol group in which the increase was of only + 12.9%, statistically insignificant.

After 5 weeks of treatment, the increase was of 65.5% in the rabbits treated with the medicine of the invention, example 1, and of 57.3% in the rabbits treated with diethylaminoethanol. Both increases were statistically significant.

Therefore, treatment with the medicine of the invention, example 1, acts faster and more intensely than DEAE on the increase of the circulating lymphocyte count.

Test 2:

The antiinflammatory action of the medicine of the invention, example 2, was studied in comparison with the action of diethylaminoethanol and diclofenac, in rats.

Inflammation was induced by intraplantar injection with carrageen, and the inflammatory oedema was measured with an Ugo Basile electronic plethysmometer.

The experiment was performed in 4 groups of male, Wistar rats weighing 120-140 g, given intraplantar injections with carrageenan, 0.1 ml from a solution 1% : 1) controls (not treated); 2) rats treated with diclofenac - 20 mg/kg bw given i.m. only once, one hour before the injection; 3) rats treated with the medicine of the invention, example 2, applied in the form of 4% gel on the paw in which carrageenan was injected, starting one hour before the carrageen injection, and then every hour for an interval of six hours; 4) rats treated with diethylaminoethanol applied locally in the form of 4% gel, using the same procedure as in the case of the group 3 rats.

The effects produced by the applied treatment were assessed by comparing the evolution of the data obtained from the measuring for 6 hours of the paw injected with carrageenan with the data obtained from the non-treated rats (controls). The groups were compared after calculating the averages and the standard deviation (Table 1).

Table 1 : Volume of the rat paw after carrageen injection in control and in treated rats, expressed as percentage of the initial level

The results have shown that, when compared to the non-treated rats, the strongest antiinflammatory effect was produced by the medicine of the invention, example 2, i.e. 72.2%.

This was followed by diehylaminoetbanol, with 56.6%, and diclofenac, with 45%.

Test 3:

The antioxidant action of the medicine of the invention, example 2, was studied in comparison with that of cfiethylaminoethanoL One tested the ability of pofymorphonuclear leukocytes (PMN) to synthesize and release oxygen free radicals (OFR).

The study was performed:

• in vivo, in PMN cells harvested from New Zealand rabbits, weighing 2000 g. They were divided into 3 groups: 1) controls, 2) injected i.m. with 25 mgkg bw of the medicine of the invention, example 2, every day, for 35 days; 3) injected i.m. with 25 mg/kg bw of diethylaminoethanol, daily, for 35 days.

• in vitro, in PMN cells harvested from healthy, non-treated human subjects. The medicine of the invention, example 2, and ckethylaπmoetbanol were added into the culture medium in an amount of 10 μg ml.

The release of oxygen free radicals by the PMN leukocytes as assessed by the chemolurninescence test

Stimulation of oxygen free radicals _^release was performed with 0.1 ml (10 mg/ml) of zimozan opsonizate (ZO) or with 0.1 ml (10 mg/ml) of concanavalinA (ConA) a) In in vivo researches, performed in PMN leucocytes harvested from treated rabbits, the results are shown in table 1.

One finds that in the rabbits treated with the medicine of the invention, example 2, the OFR level has decreased so much that, for all practical purposes, one can say the ability of the PMN cells to synthesize and release OFR has been blocked. In the rabbits treated with diethylaminoethanol, there is a tendency of OFR decrease which is not statistically significant, in the case of the noii-stimulated cells. The effect becomes statistically significant only in the case of cells stimulated with ConA, but it does not reach the level recorded in the rabbits treated with the medicine of the invention. b) In in vitro research performed in PMN leucocytes harvested from non-treated human subjects, the introduction of the medicine in cell cultures stimulated with zimozan opsonizate (ZO) induced a decrease of their capacity to release OFR in 12 of the 15 cases. The average maximum values for the entire group of investigated subjects are shown in Table 2.

Table 2: Average maximum values of FOR released by human PMN leucocytes in culture

*the medicine of the invention

The results obtained in the entire group of human subjects indicate that in the presence of the medicine of the invention, example 2, in the culture medium, the maximum level of OFR released by the human PMN leucocytes under the influence of ZO stimulator decreased by 27.4%, a statistically significant difference (p<0.01), compared to non-treated cultures.

In the presence of diethylaminoethanol, the maximum level of OFR released decreased by only 17.4%, the difference versus controls being statistically non-significant.

In conclusion, research performed in vivo and in vitro have shown that both the medicine of the invention, example 2, and diethylaminoethanol can decrease FOR synthesis and release, but the effect of the medicine of the invention is much stronger.

Test 4:

The influence of the medicine of the invention, example 1, on lymphocyte proliferation/differentiation was studied in comparison with that of diethylaminoethanol.

The labelled thymidine test has been used.

The study was performed in lymphocytes harvested from 10 adult, healthy human subjects.

It was found that addition of the medicine of the invention, example 1, in the lymphocyte culture medium together with phytoliemaglutinin (PHA) polyclonal stimulator, resulted in a greater increase of lymphocyte proliferation/differentiation than the one induced by PHA alone, while diethylaminoethanol had no influence whatsoever (Table 1). Table 1 : Stimulation indices resulting from the introduction of PHA together with the medicine of the invention, example 1, or together with diethylaminoethanol.

Therefore, the medicine of the invention, example 1, can increase by approximately 50% the stimulatory effect normally produced by PHA, unlike diethylaminoethanol that has no influence on the polyclonal effect of PHA.

Test 5:

We studied the effect of the treatment with the medicine of the invention, example 2, compared to that of dietiiylaminoethanol on the level of serum antinuclear autoantibodies in rats in which arthritis had been induced with Freund adjuvant.

Research was performed in 3 groups of 14 rats each (female Lewis rats aged 6 weeks): 1) non-treated arthritic rats (controls); 2) arthritic rats treated with the medicine of the invention, example 2, given in i.m. injections of 10 mg/kg bw and local applications of 4% gel on the extremities, two times every day for 30 days; 3) arthritic rats treated with diethylaminoethanol, administered in the same manner and using the same doses as in the case of the medicine of the invention.

After 30 days from inducing arthritis, blood was harvested from all animals and serum antinuclear autoantibodies were deteπnined by imunofluorescence.

The results are shown in the table below: Table: Level of serum antinuclear auto antibodies in the animals with arthritis induced with

Freund adjuvant.

* medicine of the invention, example 2

In the arthritic animals treated with the medicine of the invention, the serum antinuclear auto antibodies titre dropped by 33% compared to the level in the non-treated arthritic rats or to that in arthritic rats treated with diethylaminoethanol. One should note that the drop in the level of serum antinuclear autoantibodies in the animals treated with the medicineof the invention, example 2, occurred in the presence of a particularly virulent inflammatory process, such as the one produced by the Freund adjuvant.

Test 6:

One studied the effect of the treatment with the medicine of the invention, example 1, or with diethylaminoethanol on the migration of macrophages and of spleenic PMN granulocytes, and implicitly on the degree of cytotoxicity and antigenity.

Research was conducted in 3 groups of New Zealand rabbits, weighing 1800-2000 g: 1) controls, 2) rabbits treated for 30 days with 25 mg/kg bw of the medicine of the invention, example 1; 3) rabbits treated for 30 days with 25 mg/kg bw of dietiiylaminoethanol.

Macrophages and PMN granulocytes were harvested from the spleen of animals after sacrification.

We determined the macrophages migration inliibition indice (MMH), calculated as the ratio between the migration area of cells incubated for 24 hours in RPMI medium supplemented with the medicine of the invention, example 1, or with diethylaminoethanol, 20 μg/ml of each.

The results are shown in the following table 1. Table 1: Average values of migration index in macrophages and PMN leucocytes obtained from the spleen of rabbits

*= medicine of the invention, example 1

As shown in the table 1, the cells were not killed, but actually moved even more actively in the presence of the medicine which proves that the medicine of the invention is not cytotoxic or antigenic.

In the case of cells harvested from animals treated with diethylaminoethanol, addition of the same substance in the culture medium does not kill the cells, but their movement is slower than that of cells treated with the medicine of the invention, example 1.

Test 7:

The acute toxicity of the medicine of the invention, example 1, was studied by detemiining LD₅o, with the medicine given orally to rats. This was compared with the LD₅0 of dietltylaminoethanol, which is known to be of 1300 mg/kg bw. Reseai'ch was perforated in male Wistar rats, weighing 100 ± 10 g. The animals were healthy and were maintained in similar housing and feeding conditions throughout the duration of the experiment. The medicine was dissolved in distilled water and was administered by gavage feeding, in the morning, after fasting.

The animals' state of health, behaviour and mortality were monitored for 15 days after administration of the product.

The M.W. method was used to determine acute toxicity.

We found that on oral administration of the medicine of the invention, LD₅o is 9000 mg/kg bw.

The toxicity of the medicine of the invention, example 1, is therefore much lower that that of diethylaminoethanol, in which LD50 after oral administration is 1300 mg/kg bw.

One can state that the medicine of the invention, example 1, is among the substances with a very low toxicity level.

Claims

Claims:

1. Medicine for the prevention and treatment of inflammatory and degenerative diseases, characterized by the fact that it contains as active ingredient an ester of chethylaminoethanol with the phthalic or aminophthalic acid whose formula is shown in (1), or a pharmaceutically- acceptable salt thereof:

COOCH₂CH2 (C2H5)2 (I)

COOCH₂CH₂N(C₂H₅)₂

where X is H or an -NH2 group, or a pharmaceutically-acceptable salt thereof

2. Medicine, according to claim 1, characterized in that, the said medicine is formulated in the form of injectable solutions, capsules, tablets, dragees, drops, gel, cream, liniment, lotion, syrup, powder, suppositories, solutions and other pharmaceutically-acceptable forms.

3. Medicine, according to claims 1 to 2, cliaracterized in that, the said medicine is used: - in the prevention and treatment of inflammatory or chronic degenerative diseases, of immune failure and of pathological conditions with an increased production of free oxygen radicals, auto antibodies and immune complexes, such as: arteriosclerosis, cancer, AIDS; - in the treatment of inflammatory diseases of the airways, of skin and mucosa inflatnmation and of inflamed venous stasis, such as: bronchitis, rhino-pharyngitis, angina, aphtae, gingivitis, stomatitis, otitis, piodermal infections, eczema, herpes, irritations, itchings, superficial phlebitis, hemorrhoids; - in the treatment of rheumatic processes, such as: rheumatoid arthritis, arthritis and arthrosis, lombosciatics, tendinitis, myositis, rachialgia, neuralgia, myalgia;

4. Use of the medicine according to claim 1 for the prevention and treatment of inflammatory and degenerative diseases characterized in that it consists in administering the medicine according to claims 1 to 3: - in the form of capsules, tablets, dragees, ampoules and the like for immune system stimulation, nootropic effect and treatment of chronic degenerative diseases; - in the form of capsules, dragees, tablets, ampoules, gel, solution, cream, ointment and the like for the treatment of rheumatoid arthritis and of other rheumatic processes; - in the form of lotion, gel, cream, ointment, powder, drops, suppositories and the like for the treatment of skin and mucosa inflammation; - in the foπn of capsules, dragees, tablets, syrup, ampoules, aerosols solution and the like for the treatment of airways inflammation. The medicine is aiiministered in doses of 5...150 mg kg bw/day, preferably 30 mg/kg bw/day, given orally, in series of 15-30 days, with pauses of 15-45 days, for prevention purposes, and of 10...100 mg/kg bw/day, preferably 30 mg/kg bw/day, given parenterally, or in doses of 15...200 mg/kg bw/day, preferably 60 mg/kg bw/day, given orally and/or in concentration of 4...50%, preferably 10 g%, in local applications, for 3-20 days, for curative purposes.