RU2144918C1 - 3-benzylaminomethylenepyrrolidine-2,4-dione and anticonvulsant agent based on said - Google Patents

Abstract

FIELD: organic chemistry, pharmacy. SUBSTANCE: invention relates to the new chemical compound - 3-benzylaminomethylenepyrrolidine-2,4-dione. Compound shows the more expressed anticonvulsant activity, broad spectrum of action and decreased toxicity as compared with the known anticonvulsant preparation widely used in clinics. Invention describes also the drug eliciting anticonvulsant effect. EFFECT: new compound indicated above, improved properties of compound. 3 cl, 12 tbl, 1 ex

Description

и лекарственного препарата противосудорожного действия на его основе.The invention relates to organic chemistry and medicine and relates to a new chemical compound, namely a derivative of pyrrolidine-2,4-dione of the formula I:

and an anticonvulsant drug based on it.

где R - атом фтора, хлора и другие, обладающие трнаквилизирующей и антиконвульсивной активностью. (Акцептованная заявка Японии N 73-34741, C 07 D 27/06, A 61 K 27/00, 23.10.73 г.)
Однако в описании данной заявки отсутствуют какие-либо количественные характеристики биологической активности, что не позволяет провести сопоставительный анализ заявляемого соединения с соединениями Японской заявки.The literature describes pyrrolidinedione general formula II:

where R is an atom of fluorine, chlorine, and others with trinquilizing and anticonvulsant activity. (Japanese Accepted Application N 73-34741, C 07 D 27/06, A 61 K 27/00, 10/23/73)
However, in the description of this application there are no quantitative characteristics of biological activity, which does not allow for a comparative analysis of the claimed compounds with compounds of the Japanese application.

которая является нейропередатчиком во вставочных ГАМК-эргических нейронах и выполняет тормозные функции в центральной нервной системе (Davidoff R. A. , in: Amino acid neurotransmitters (Ed. by F.V. De Fendis, P. Mandel), N. V. , Raven Press, 1981, p. 249 - 255. Fagg G.E., Forster A.C., Neuroscience, 1983, v. 9(4), p. 701 - 719)
ГАМК принимает участие в качестве одного из главных тормозных медиаторов в регуляции моторной активности, поддержании судорожного порога, формировании эмоционального поведения, контроле высвобождения ряда гипофизарных гормонов, осуществлении высших интегративных функций мозга, таких как условные рефлексы, процессы обучения и памяти.The structure of compound 1 contains elements characteristic of γ-aminobutyric acid (GABA) of formula A:

which is a neurotransmitter in intercalated GABAergic neurons and performs inhibitory functions in the central nervous system (Davidoff RA, in: Amino acid neurotransmitters (Ed. by FV De Fendis, P. Mandel), NV, Raven Press, 1981, p. 249 - 255. Fagg GE, Forster AC, Neuroscience, 1983, v. 9 (4), p. 701 - 719)
GABA takes part as one of the main inhibitory mediators in the regulation of motor activity, maintaining the seizure threshold, the formation of emotional behavior, controlling the release of a number of pituitary hormones, and the implementation of higher integrative brain functions, such as conditioned reflexes, learning and memory processes.

 (K.S. Raevsky, V.P. Georgiev, in the book: Mediator amino acids: neuropharmacological and neurochemical aspects - M., 1986, p. 55-56).

 In this regard, the anticonvulsant activity of the claimed compounds should be compared with analogues in action - with anticonvulsant drugs: sodium valproate, the mechanism of action of which is due to the accumulation of GABA in the brain as a result of inhibition of the enzyme GABA-transaminase, and diphenin, which is widely used in the clinic of epilepsy against large convulsive seizures (M.D. Mashkovsky, in the book: Medicines. - Minsk: Publishing House "Belarus", 1987, p. 142 - 154.

Sodium valproate - sodium salt (2-propylvaleric acid) or dipropylacetic acid of the formula III:
(CH ₃ CH ₂ CH ₂ ) ₂ CHCOONa III (sodium valproate)
used in the clinic of epilepsy for large and small seizures (grand et pelit mol) seizures (M. D. Mashkovsky, in the book: Medicines. - Minsk: Publishing House "Belarus", 1987, p. 142 - 154). According to experimental data, it has an anticonvulsant effect on many models of convulsive states (convulsions caused by MES, perezol, GABA-deficient convulsions), but in fairly large doses, approximately 1/3 of LD ₅₀ . In this regard, the therapeutic doses used in the clinic are also high (0.25 g - 1 g). The drug is practically inactive for seizures caused by cholinomimetic substances.

в эксперименте обладает выраженным влиянием на тоникоэкстензорные судороги, вызванные максимальным электрошоком (МЭШ) и стрихнином. Оказывает слабый эффект на моделях судорог, вызванных аудиогенным раздражителем, практически неактивен при ГАМК-дефицитных судорогах и судорогах, вызванных коразолом и холиномиметическими веществами. По токсичности относится к веществам со средней токсичностью (Masuda V., Karasava T., Shiraishi V., et al. , Arzneim. Forsch / Drug. Res., 1980, v. 30(1), p. 477 - 483).By toxicity, sodium valproate belongs to low-toxic substances (Loscher W., Meldrum BS, Federation Proc, 1984, v. 43, p. 276 - 284)
The drug diphenin (5,5-diphenylhydantoin-sodium) - formula IV:

in the experiment, it has a pronounced effect on tonic-extensor convulsions caused by maximum electroshock (MES) and strychnine. It has a weak effect on models of seizures caused by an audiogenic stimulus and is practically inactive in GABA-deficient seizures and seizures caused by corazole and cholinomimetic substances. By toxicity, it refers to substances with moderate toxicity (Masuda V., Karasava T., Shiraishi V., et al., Arzneim. Forsch / Drug. Res., 1980, v. 30 (1), p. 477 - 483).

 The purpose of the invention is a new derivative of a series of pyrrolidin-2,4-dione, which has a more effective anticonvulsant activity, breadth of therapeutic effect, low toxicity.

 The goal is achieved by the chemical structure of the claimed compounds 3-benzylaminomethylene pyrrolidine-2,4-dione.

 The inventive compound has anticonvulsant activity. Anticonvulsant drugs, consisting of the active substance and a pharmaceutical carrier, according to the invention, as the active substance contains the inventive compound - 3-benzylaminomethylene pyrrolidine-2,4-dione of the above formula I.

 The inventive preparation can be used orally in various dosage forms (dragees, capsules, suppositories).

 Preferably, according to the invention, the inventive preparation is used in the form of tablets with an active substance content of 0.1 to 0.25 g per tablet.

 The inventive compound is a white crystalline substance, stable in air, soluble in chloroform, dimethylformamide, dimethyl sulfoxide, sparingly soluble in water and alcohols.

The structure of compound 1 is proved by spectral data and is confirmed by the data of elemental analysis, mass spectrum, which contains the peak of the molecular ion M ⁺ 216 and the peaks of m / e 199 ions (M ⁺ -OH), m / e 159 (M ⁺ -COCH ₂ NH ), m / e 91 (PhCH ₂ ).

In the IR spectrum, absorption bands are observed at 3180 and 3400 cm ^-1 (NH-group), 1695 cm ^-1 (CO-ketone), 1645 cm ^-1 (CO-lactane), 1605 cm ^-1 (C = C).

According to NMR ¹ H-spectra (d-DMSO), the claimed compound is a mixture of cis-trans isomers relative to the C = C bond. The spectrum contains ppm signals (d ₆ -DMSO): 3.54 and 3.57 (doublets J ≈ 1 Hz, 5-CH ₂ ), 4.57 and 4.61 (doublets, J ≈ 3 Hz, PhCH ₂ ), 7.48 (broad, 1-NH), 7.30 - 7.39 (m, Ph), 7.69 and 7.90 (doublets, J ≈ 15 Hz) (CH), 9 42 and 9.49 (m, NH-CH ₂ ). The ratio of isomers upon crystallization from various solvents (water, MeCN, MeOH) ≈ 1: 1.

¹³ C NMR spectrum: 49.6 and 50.5 (PhNHCH ₂ ), 52.0 and 52.3 (5-CH ₂ ), 127.8; 127.9; 128.8 and 140.0 (Ph), 140.0 (= CH), 98.2 and 97.2 (C ³ ); 152.5 and 152.3 (2-CO); 171.6 and 173.7 (4-CO) ppm. (d ₆ -DMSO).

 The method of obtaining the claimed compound is based on the well-known transamination reaction.

с бензиламином в среде полярного растворителя (предпочтительно низшего спирта) в температурном интервале 5 - 80^oC (предпочтительно при комнатной температуре).3-Benzylaminomethylenepyrrolidine-2,4-dione is obtained by the interaction of 3-dimethylaminomethylenepyrrolidine-2,4-dione (IV):

with benzylamine in a polar solvent (preferably lower alcohol) in the temperature range of 5 - 80 ^o C (preferably at room temperature).

 Starting compound IV was prepared by reacting diethyl acetal dimethylformamide with pyrrolidine-2,4-dione.

 The following example illustrates the preparation of the inventive compound 3-benzylaminomethylene pyrrolidine-2,4-dione.

 To a solution of 15.4 g (0.1 m) of 3-dimethylaminomethylene pyrrolidin-2,4-dione in 150 ml of methanol was added 12.8 g (0.13 m) of benzylamine, the mixture was kept at room temperature for 1 hour, then cooled. The precipitated crystals are filtered off, washed with methanol and dried.

19.0 g (88%) of 3-benzylaminomethylene pyrrolidin-2,4-dione are obtained, mp. 165-167 ^° C (CH ₃ OH).

Calculated,%: C 66.65 H 5.59 N 12.96
C ₁₂ H ₁₂ N ₂ O ₂
Found,%: C 66.87 H 5.72 N 12.95 M ⁺ 216.

 The inventive compound and a drug based on it were tested in an animal experiment.

 A study of the biological activity of the claimed compounds was carried out with respect to control and anticonvulsants: sodium valproate, diphenin. The results of the study are presented in table. from 1 to 12.

 As a result of the studies, it was found that the claimed compound 1 has a pronounced anticonvulsant activity in most models of convulsive conditions and has a minimal side neurotoxic effect.

 As can be seen from the table. 1 and 2, the claimed compound by its effect on tonic convulsions in mice and rats caused by maximum electroshock (MES) is not inferior to diphenin (ED 50 for 1 is 26 mg / kg for diphenin is 24 mg / kg) and exceeds sodium valproate in 20 times.

 The therapeutic breadth of the claimed compounds is 30 times greater than that of sodium valproate and 6 times greater than diphenin.

 The claimed compound 1, like diphenin, does not prevent the clonic component of convulsions caused by corazole, but is 3.5 times greater than valproate for the prevention of the tonic phase of seizures (Table 3) and has a pronounced effect on the generalization of convulsive activity caused by corazole in rat cerebral cortex (table 4). At a dose 2 times lower (60 mg / kg) than sodium valproate (120 mg / kg), compound 1 prevents the development of corazole "Kindling" in rats, while diphenin, even at the maximum dose (100 mg / kg), practically does not affects the effect of corazole "Kindling" (table 5).

 A study of the GABAergic link in the anticonvulsant mechanism showed that the claimed compound prevents seizures and death of animals caused by the GABA receptor antagonist bicucullin - in doses comparable to doses of sodium valproate, while diphenin practically does not show such an effect (Table 6 ) In addition, in experiments with the convulsive effect of d, l-allyl glycine on rats, it was shown that compound 1 effectively prevents the development of tonic seizures (ED 50 = 51 mg / kg) and, in particular, their tonic-extensor phase (ED 50 = 13 mg / kg).

 Compound 1 also prevents cramps in mice caused by strychnine (ED 50 for 1 is 155 (115-175 mg / kg, oral), while reference drugs do not.

 According to the effect on the convulsive effect of cholinomimetic substances (nicotine and arecoline), compound 1, like sodium valproate, and diphenin, has an insignificant effect (somewhat shortening the latent period of the onset of seizures, Table 7).

 In the model of audiogenic epilepsy in rats, the claimed compound (40-400 mg / kg, by mouth) causes a weakening of the audiogenic epileptiform reaction (increase in the latent period, decrease in the intensity of seizures) and in 100% of cases removes the tonic phase of the seizure and increases the latent period of the onset of motor excitation. According to the effect on the latent period and seizure intensity, the claimed compound is comparable with valproate after administering them after an hour, but after 24 hours this effect is lost in valproate, while it remains in the claimed compound and continues to persist for 7 days (table 8). In these experiments, diphenin (2 and 20 mg / kg, by mouth) practically does not affect the severity of the audiogenic reaction.

 The claimed compound 1 in doses of 100 and 200 mg / kg, like diphenin and sodium valproate, has anticonvulsant activity on the model of focal epileptiform activity in the rat cerebral cortex, causing a sharp decrease in the total number of electroconvulsive discharges against a background of a certain increase in the frequency of peak discharges, which characteristic of anticonvulsants.

 The study of adverse neurotoxic effects showed that the claimed compound as well as comparison drugs have a depressing effect on the central nervous system, accompanied by a coordination disorder and a hypnotic effect (Table 10). However, these effects are observed at doses ten times greater than the effective doses of the anticonvulsant effect (Tables 1 and 8). The ratio of doses that cause a neurotoxic effect (impaired coordination) with doses that have an anticonvulsant effect (convulsions caused by MES) showed that the claimed compound 1 has a higher therapeutic latitude index than sodium valproate and diphenin (Table 1).

 According to the general pharmacological action, the claimed compound 1 in a therapeutic dose does not inhibit spontaneous motor activity in mice and does not inhibit the development of a conditioned reflex of active avoidance in rats, while sodium valproate has a depressing effect on these parameters. In addition, the claimed compound has an anti-aggressive effect in experiments on mice with clonidine and pain-provoked aggression and is not inferior to sodium valproate (Tables 11 and 12).

Compound 1, like sodium valproate, is characterized by low toxicity (their LD ₅₀ ) is 2400 and 2068 mg / kg, by mouth), while diphenin toxicity is 363 mg / kg, by mouth (Table 1). In in vitro tests on microsomal objects of liver cells, pig testes cells and mitochondrial membranes of bovine adrenal glands, compound 1 does not exhibit the property of an enzyme inducer of the P-450 system.

 Thus, the above experimental data suggest that the claimed compound is 3-benzylaminomethylene pyrrolidine-2,4-dione-a new compound of the pyrrolidine-2,4-dione series, has pronounced anticonvulsant properties, minimal side neurotoxic effects and low toxicity. The claimed compound has distinct advantages compared with the comparison drugs (sodium valproate and diphenin), in most tests characterizing anticonvulsant activity, as well as in terms of adverse neurotoxic effects and acute toxicity.

 In this regard, the claimed compound is of interest as an active substance of an anticonvulsant (antiepileptic) agent, which can be used in various dosage forms (even capsules, suppositories).

 Preferably, the inventive preparation is used in the form of tablets with an active substance content of 0.1-0.25 g per tablet.

A pharmacological study on animal tablets containing compound 1 of the following composition was carried out:
Compound 1 - 0.25 g
Microcrystalline cellulose - 0.06 g
Milk sugar - 0.08 g
Potato starch - 0.105 g
Calcium stearate - 0.005 g
The average weight of one tablet is 0.5 g
The number of tablets - 100 pcs.

1. Effect on seizures caused by maximum electroshock in mice and rats
The study of tablets of this composition was carried out in comparison with the substance of compound 1 according to some indicators of anticonvulsant action and indicators characterizing neurological disorders in mice and rats.

 The anticonvulsant effect was studied in experiments on white male mice (weighing 18–20 g) and male rats (weighing 100–120 g); seizures were caused by the ECT UNIT constant pulse generator (Ugo Basile, Italy). An electric discharge with parameters of 50 mA, 60 Hz, 2 sec in mice was passed through corneal electrodes. In rats, MES was induced by passing current through the ear electrodes with parameters: 100 mA, 80 Hz, 0.2 sec. Seizures were characterized by tonic extension of the hind limbs and ended in the death of 60-80% of the animals in the control groups.

 The study was carried out in 4-5 doses (in groups of 10-20 animals) (a total of 120 mice and 80 rats were used), which were administered orally as a suspension in a 1% solution of carboxymethyl cellulose (CMC) in a volume of 0.3 ml for mice and 0 , 5 ml - for rats 2 hours before the application of maximum electroshock (MES).

 In the experimental and control groups, the number of animals was recorded in which there was a tonic-extensor reaction after applying MES.

 The severity of the anticonvulsant effect of the studied objects was evaluated when calculating the effective doses (mg / kg) preventing the development of a convulsive reaction in 50% of animals (ED 50).

 In mice, compound 1 (tablets and substance) was administered in suspension in a 1% CMC solution orally in the dose range of 12.5-100 mg / kg for 1 hour MES. Anticonvulsant activity was evaluated by the prevention of tonic-extensor seizures in 50% of animals, the intensity of development and the duration of the anticonvulsant effect. The ED of 50 studied drugs was calculated.

 Studies have shown that in mice, compound 1 in the form of tablets has an anticonvulsant effect and does not significantly differ in effectiveness from substance (Table 13). The anticonvulsant effect of the tablets and substance was well manifested after 30 minutes and reached a maximum after 1-1.5 hours.

 In rats, the anticonvulsant activity of the tablets was also evaluated by preventing the tonic-extensor component, the intensity of development and the duration of the anticonvulsant effect. The severity of the anticonvulsant effect was judged on the basis of the values of effective doses (ED 50) calculated for tablets and substance.

 Studies have shown that in rats, the tablets of compound 1 have an anticonvulsant effect and do not significantly differ in effectiveness from the substance (Table 14).

Effect on seizures caused by corazole in mice
The effect of compound 1 in the form of tablets and substance on the convulsive effect of corazole was studied on male mice weighing 20-22 g.

 When studying the effect on the conical and tonic-extensor component of convulsions, corazole was administered to mice at a dose of 175 mg / kg, intraperitoneally. In this case, the convulsions had a mixed clonic-tonic character and ended in the death of animals at the height of the tonic-extensor phase. Effective doses (ED 50) were determined that prevented the tonic-extensor component of fore and hind limb cramps in 50% of the animals.

 Studies have shown that tablets, like substance, do not affect the clonic phase, but in a dose-dependent manner they prevent the development of the tonic-extensor phase of corazole seizures (Table 15). According to the activity of the tablets, it does not significantly differ from compound 1 in the form of a substance.

Neurological Disorders in Mice (Imbalance)
Evaluation of neurological disorders was performed in mice in the test "rotating rod". The presence of drugs with a similar effect was judged by the number of animals that were unable to stay on the rod for 2 minutes.

 In these experiments, the tablets and substance of compound 1 were used in a dose range of 100-1500 mg / kg, inside as a suspension in a 1% CMC solution, administered 1 hour before placing the animals in the device.

 Studies have shown that the administration of tablets or substances of compound 1 in doses exceeding the therapeutic ones due to the anticonvulsant effect (MES-convulsions) caused a dose-dependent disorder of coordination of movements in animals. According to this indicator, the tablets of compound 1 also did not differ significantly from the substance (Table 16).

 Thus, studies have shown that compound 1 in the form of tablets in experiments on mice and rats has anticonvulsant activity and does not significantly differ in activity from substance.

 The test results confirmed the possibility of drug transfer to non-clinical trials.

Claims (1)

1. 3-Benzylaminomethylenepyrrolidine-2,4-dione of the formula I

2. An anticonvulsant, characterized in that its active substance is 3-benzylaminomethylene pyrrolidine-2,4-dione according to claim 1.

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