RU2554502C9 - Medication for treating cerebral circulation disorders and brain traumas - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: claimed is the application of N-(2-adamantyl)-hexamethylenimine hydrochloride (himantane) as a medication for treating cerebral circulation disorders and brain traumas. An ability of himantane to reduce animal morbidity and increase cognitive and motor indices in case of its introduction after 3.5 hours after modelling a hematoma in rats and further introduction for 4 days was determined, which testifies to its possessing the neuroprotective potential on the model of the intracerebral post-traumatic hematoma. The technical result consists in the realisation of the claimed purpose: it has been shown that himantane in a dose of 5 mg/kg in case of intravenous introduction causes the increase of cerebral circulation in the rats' brain after ischemic brain affection without changing the level of arterial pressure.

EFFECT: medication did not produce a significant impact on the blood supply of the brain of the intact animals and did not cause a change in the heart rate and respiratory rate.

1 dwg, 7 tbl

Description

The invention relates to medicine, in particular to pharmacology, and relates to a new application of the known substance - N- (2-adamantyl) -hexamethyleneimine hydrochloride (gimantan) (patent No. 1825499) - for the treatment of cerebrovascular disorders and brain injuries.

Parkinson's disease is a neurodegenerative disease characterized by progressive death of dopaminergic neurons. For the pathogenetic therapy of this disease, the search for drugs that slow and prevent the neurodegenerative process is relevant. Neurodegeneration of brain tissue is also the result of strokes, mortality and disability in which is an acute problem for modern health care. The neuroprotective drugs used for the treatment of neurodegenerative diseases and, in particular, Parkinson's disease, are promisingly considered as potential drugs for the treatment of cerebrovascular disorders and brain injuries.

The present invention describes a new use of the antiparkinsonian drug Himantan as an agent for the treatment of cerebrovascular accidents and brain injuries.

The prototype of the present invention is a low-affinity non-competitive blocker of the ion channel of the glutamate receptors NMDA subtype amantadine, which is widely used to attenuate motor disorders in the early stages of Parkinson's disease, due to the neuroprotective properties of the injection form which is used for traumatic brain injury and acute cerebrovascular accident. It was shown that in case of traumatic brain injuries, the administration of amantadine at a dose of 200-400 mg per day caused an improvement in memory and attention in patients (E. Sawyer, L.S. Mauro, M.J. Ohiinger Ann Pharmacother 2008; 42 (2): 247-252). The administration of amantadine hydrochloride for 4 weeks to patients in a post-traumatic vegetative state or with minimal consciousness accelerated functional recovery, but when the drug was discontinued, a deterioration was noted in comparison with patients who received a placebo (D. Brunk. Clin neurol news 2011; 7 (6): 12- 13).

The administration of amantadine sulfate in the acute period of ischemic stroke led to a significant restoration of the level of consciousness and improvement in the dynamics of regression of neurological disorders with the most intensive recovery of neurological deficiency on the 1st day (D.R.Khasanova, R.X. Khafizyanova, R.R.Islamov and et al. Journal of Neurology and Psychiatry named after S.S. Korsakov 2009; 5: 37-43). The use of the drug daily intravenously for 10 days in patients in the acute period of ischemic stroke accelerated the recovery of motor disorders and reduced spasticity (E.I. Gusev, G.N. Avakyan, E.A. Katunina and others. Zh. Neurology and psychiatry. S.S. Korsakova, Stroke. 2006; 18: 55-57).

In experimental studies, it was found that gimantan in doses of 5-20 mg / kg has a wide spectrum of antiparkinsonian activity: it attenuates the main manifestations of experimental parkinsonism syndrome in C57BL / 6 mice and rats - oligokinesia, rigidity, tremor. Gimantan as an anti-Parkinsonian drug is superior to amantadine in the dose range and spectrum of specific activity. A significant advantage of gimantan is the ability to eliminate tremor. Gimantan, unlike amantadine, has an anxiolytic and antidepressant effect (E.A. Waldman. Author. Abstract. Thesis. Dr. med. Sciences, 2001, Moscow). The main neurochemical mechanisms of action of gimantan have been established: low-affinity non-competitive blockade of the ion channel of the glutamate NMDA receptor subtype at concentrations achieved with systemic administration (M.V. Elshanskaya, A.I. Sobolevsky et al. Experimental and wedge pharmacology 2001; 64 (1) : 18-21); noncompetitive reversible inhibition of monoamine oxidase B (MAO-B) (E.A. Waldman, T.A. Voronina, L.I. Aksenova et al. Experimental and wedge pharmacology 2003; 5: 3-5). Gimantan in a therapeutic dose increases the content of extracellular dopamine and reduces the content of its metabolites in the rat striatum. Gimantan has a modulating effect on subtypes of dopamine and serotonin receptors, as well as on the reuptake of dopamine. Subchronic administration of hematan to rats ex vivo leads to an increase in the number of D ₁ receptors in the striatum and 5-HT _1A receptors in the hippocampus with a simultaneous decrease in the number of 5-HT _2A receptors in the frontal cortex, both in normal and in case of MPP + - induced Parkinson's syndrome in animals. The acute administration of gimantan does not affect the level of the dopamine transporter protein (DAT) in the striatum of C57B1 / 6 mice, as well as the rate of reuptake of [3H] -DA; co-administration of the drug with MPTP causes a significant decrease in DAT. Subchronic administration of gimantan to intact animals and animals with a toxin background, in contrast, leads to an increase in the amount of DAT in the striatum and frontal cortex, and also increases the rate of reuptake of [3H] -DA in the striatum (G.I. Kovalev, D.A. Abaimov, M.V. Voronin et al. Neurochemistry 2007; 24 (2): 150-156; D.A. Abaimov, G.I. Kovalev. Experimental and wedge pharmacology 2007; 70 (4): 11-14). Gimantan exhibits antioxidant activity in in vitro and in vivo model systems. Gimantan does not adversely affect learning and memory, does not have addictive potential, does not alter the psychomotor activity of animals, that is, it does not have side effects inherent in some NMDA receptor antagonists (E.A. Waldman. Author. , 2001, Moscow).

Direct and indirect via NMDA receptors effect on dopamine receptors, release, reuptake and metabolism of dopamine contribute to an increase in dopaminergic transmission activity and cause the symptomatic effect of the drug in Parkinson's disease. In a clinical study, gimantan effectively eliminated major motor disorders in patients with early stages of Parkinson's disease when applied at a dose of 25 mg per day (E.A. Katunina, A.V. Petrukhova, G.N. Avakyan, et al. Journal of Neurology and Psychiatry named after S.S. Korsakov 2008; 108 (6): 24-27).

The complex mechanism of action, including the properties of a low-affinity non-competitive blocker of the ion channel of the glutamate NMDA receptor subtype, a non-competitive inhibitor of MAO-B, a modulator of subtypes of dopamine and serotonin receptors and reuptake of dopamine, antioxidant properties suggest the presence of a neuroprotective agent in hematan and its pathogen therapy and its activity Parkinson's, since glutamate excitotoxicity, oxidative stress and neuroinflammation are ove advantageously progressive degeneration of dopaminergic neurons in the disease. However, these same processes underlie the death of neurons in other acute conditions, in particular in cases of cerebrovascular accidents and brain injuries.

The essence of the present invention is to identify the possibility of using gimantan for the treatment of brain injuries and cerebrovascular accidents and is illustrated by the following examples.

Example 1

The effect of gimantan on the dynamics of disorders caused by intracerebral post-traumatic hematoma, with different methods of administration

The experiments were carried out on white non-linear male rats weighing 220-250 g, obtained from the closed colony of the Stolbovaya nursery RAMP.

Modeling of intracerebral post-traumatic hematoma was carried out according to the described technique (A.I. Makarenko, N.S. Kositsin, N.V. Pasikova, M.M.Svinov. Journal of Higher Nervous Activity 2002; 52 (6): 760-763; T.L. Garibova, I.P. Galaev, T.A. Voronin et al. Experimental and Wedge Pharmacology, 2003; 66 (3): 13-16). Rats were anesthetized with chloral hydrate (400 mg / kg, ip). After craniotomy using a special device (mandrin - knife) and stereotaxis, brain tissue was destroyed in the area of the inner capsule. After 2-3 minutes, 0.02-0.03 ml of blood taken from the tail vein of the animal was injected into the lesion site. In this way, a local autohemorrhagic stroke is achieved in the area of the internal capsule (diameter - 2 mm, depth - 3 mm) without significant damage to the superior formations of the brain and neocortex.

Gimantan was used at a dose of 5 mg / kg as an injectable dosage form (50 mg in 2 ml) for intravenous administration and at a dose of 10 mg / kg as a substance for intraperitoneal administration. Gimantan was dissolved in physiological saline. Animals were divided into 4 groups:

1. Passive control - falsely operated animals (LO), which under anesthesia conducted only craniotomy.

2. Active control - animals with intracerebral hematoma (IG).

3. The group "Gimantan 5 mg / kg iv" - rats with IG, which were administered intravenously gimantan at a dose of 5 mg / kg.

4. The group "Gimantan 10 mg / kg ip" - rats with IG, which were administered intraperitoneally gimantan at a dose of 10 mg / kg.

The first injection of hematanum was made 3.5-4 hours after surgery, then daily once a day for four days. The most effective doses of the drug in the previous studies of antiparkinsonian activity were used with the appropriate route of administration (E. A. Waldman. Author. Abstract. Doctor of Medical Sciences, 2001, Moscow; RF patent No. 1825499; application for the grant of a patent for invention No. 2011124339 dated June 16, 2011). Animals of groups 1 and 2 were injected with physiological saline in an equivalent volume.

The dynamics of disorders caused by intracerebral post-traumatic hematoma, and the effect of hematan with different routes of administration were evaluated within 14 days with registration of changes in coordination of movements in the test "rotating rod" and muscle tone by pulling on a horizontal bar. Assessment of learning and memory was carried out in the test of the conditioned reflex of passive avoidance (passive avoidance reaction) on the first, third, seventh and fourteenth days after the operation. The survival of rats was recorded within 14 days after the operation (S.B.Seredenin, V.A. Kraineva. Exper. And wedge. Pharmacology, 2009; 72 (1): 24-28.). Statistical processing of the data was carried out using the computer programs Statistica-8 (Statistika Inc., USA) and Biostat using the Shapiro-Wilk test to assess the normality of the distribution, the Student t-test for independent samples and the Fisher test. The results were considered significant at p <0.05.

results

Animal survival

Within two weeks after surgery, rat death was observed in all experimental groups. The highest mortality was recorded in the active control group and amounted to 40% of animals, which was 30% higher compared to the group of falsely operated (LO) rats, in which 10% of the animals died. In both groups of animals with IG, which were administered intravenous and intraperitoneal administration of hematan, 20% of the rats died.

Effect on coordination and muscle tone

Violations of coordination of movements and weakening of muscle tone were expressed in most animals with IG on the 1st and 3rd days with a gradual improvement by the 14th day, when violations were recorded in 25% of the animals. In LO animals, there was a slight decrease in these indicators on the first day with a complete recovery by the 7th day of observation. In groups of animals with IG, which was administered gimantan, violations were significantly less pronounced than in untreated rats with IG. Complete recovery by the 14th day was recorded in both groups of animals treated with gimantan (table 1).

Table 1 The effect of hematan with different modes of administration on coordination and muscle tone of rats with intracerebral hematoma The proportion of animals of the group that did not stay on a rotating rod (v.) And did not catch up on a horizontal crossbar (etc.),% 1st day 3rd day 7th day 14th day Group of animals w.s. g. w.s. g. w.s. g. w.s. g. LO 25 25 5.5 5.5 0 0 0 0 IG 66.3 * 66.3 * 57.2 57.2 53.8 46.16 25 25 Gimantan 5 mg / kg iv 40 40 10# 10# eleven# eleven 0 0 Gimantan 10 mg / kg ip twenty # twenty # 33.3 33.3 0 # 12.5 0 0 * - p <0.05 compared with the group of falsely operated rats # - p <0.05 compared with the group with intracerebral hematoma (Fisher's exact test)

Memory effect

In the passive avoidance reaction test, before conducting the training session, the mink reflex was evaluated by the latent time of the first entry into the dark compartment of the installation. There were no significant differences in the groups of animals in terms of the time the reflex was performed, which indicates identical initial conditions for training.

When assessing the reproduction of the memorial trace after 24 hours, it was found that in the group of animals with IG the latent period of entry into the dark chamber was significantly lower than in LO, most animals did not remember about electric shock. When reproducing passive avoidance reaction on the 3rd, 7th, and 14th day after training, a gradual extinction of the memorial trail was observed in the group of animals, but on all days of observation, the indicator remained significantly higher than in the group of animals with IG.

The introduction of hematanum 5 mg / kg intravenously for 5 days after the operation completely prevented memory impairment in animals with IG; the indices of reproduction of the memorable trace did not differ from the LO group on all days of observation. The effectiveness of gimantan when administered intraperitoneally at a dose of 10 mg / kg was comparable to its activity when administered intravenously at a dose of 5 mg / kg (Fig. 1).

Example 2

The effect of gimantan on the blood supply to the brain of intact rats and blood supply to the brain of animals in conditions of global transient ischemia

The experimental work was carried out on white non-linear male rats weighing 250-300 g, obtained from the closed colony of the Stolbovaya nursery RAMS.

Animals were anesthetized with urethane (1200 mg / kg, ip). Gimantan was used at a dose of 5 mg / kg and 10 mg / kg as an injectable dosage form (50 mg in 2 ml) with intravenous administration. Blood flow to the brain was recorded in the carotid artery system using a T106 ultrasonic flowmeter from Transonic Systems Inc. (USA). A flowmeter sensor with a diameter of 1 mm was installed on the common carotid artery of animals after ligation of the external carotid artery. Blood pressure was recorded in the femoral artery through a silicone catheter, and the drug was administered through a silicone catheter into the femoral vein. To assess the state of microcirculation in the vessels of the brain, local cerebral blood flow was recorded in the parietal region of the rat cerebral cortex using an ALF-21 laser Doppler flowmeter from Transonic System Inc. (USA). For this purpose, a 0.8 mm diameter flowmeter needle sensor was installed on the parietal region of the cerebral cortex at a distance of 6-7 mm distal to the base of the middle cerebral artery in the direction of its central branch using a micromanipulator and rocker arm.

Global transient ischemia in rats was caused by occlusion of both common carotid arteries for 10 minutes. At the same time, the blood pressure level was reduced to 40-50 mm Hg by bloodletting. Art. A decrease in blood pressure is necessary for impaired functioning of the vertebrobasilar system, which provides blood supply to vital centers located in the brain stem region. After 10 minutes, the clamps were removed and the blood was reinfused. Gimantan was administered 40-45 minutes after global transient brain ischemia.

All the studied parameters were recorded using a polygraph company "BIOPAK" (USA), the recording of indicators was carried out on a personal computer.

Statistical data processing was performed using the Stastistika 8.0 software package (Statistika Inc., USA). To assess the normality of the distribution, the Shapiro-Wilk test was used. Since there was no normal distribution, the Wilcoxon non-parametric two-sample sign-rank test was used for further data processing. The results were considered significant at p <0.05.

results

The effect of gimantan on blood supply to the brain of rats

Experiments with anesthetized rats showed that a dose of 5 mg / kg gimantan when administered intravenously causes a slight decrease in cerebral blood flow in the internal carotid artery by an average of 5.25% (20 minute, table 2). A decrease in blood flow is observed within 60 minutes of observation, however, from 30 minutes to the end of the experiment, its decrease is statistically insignificant.

Against the background of a single intravenous administration of hematan at a dose of 5 mg / kg at 20 minutes of the experiment, there was a tendency to increase the level of blood pressure (table 3). However, already from 30 minutes to the end of the experiment, the level of blood pressure statistically slightly decreased.

table 2 The effect of hematan (5 mg / kg, intravenously) on cerebral blood flow (MK) in the carotid artery in anesthetized rats (ml / min; MK fraction relative to background,%) Background ml / min 10 min after preparation ml / min 20 min after preparation ml / min 30 min after preparation ml / min 40 min after preparation ml / min 50 min after preparation ml / min 60 min after preparation ml / min 10 min % 20 minutes % 30 minutes % 40 min % 50 min % 60 min % Minimum 33 3.2 -8.1 2.7 -23.1 2.7 -18.2 2,8 -15.2 2,8 -15.4 3.0 -13.5 25th percent 4.2 4.3 -4.4 4.0 -10.8 4.3 -10.8 4.3 -3.9 4.0 -13.5 3.8 -9.1 Median 4.95 4.9 -3.25 4,5 * -5.25 4.7 -5.6 4.95 -0.1 4.75 -3.1 4.6 -3.0 75th percent 5.7 5,4 1.9 5,4 -2.1 5,4 0,0 5,6 2,4 5,6 4.9 5.8 1.8 Maximum 6.1 6.2 11.9 6.1 2,4 5.7 9.5 6.2 8.3 6.4 6.3 6.8 11.5 * - p = 0.01 - in comparison with the background

Table 3 The effect of gimantan (5 mg / kg, intravenously) on the blood pressure (BP) of anesthetized rats (mmHg). st .; its change relative to the background,%) Background ml / min 10 min after preparation ml / min 20 min after preparation ml / min 30 min after preparation ml / min 40 min after preparation ml / min 50 min after preparation ml / min 60 min after preparation ml / min 10 min % 20 minutes % 30 minutes % 40 min % 50 min % 60 min % Minimum 77.0 81.0 -7.7 86.0 -11.5 71.0 -16.7 77.0 -25.0 78.0 -25.0 75.0 -27.9 25th percent 104.0 104.0 -3.1 102.0 0,0 92.0 -7.8 86.0 -9.8 85.0 -9.8 86.0 -6.5 Median 114.5 116.5 4.4 123.5 3,5 107.5 0.35 113.0 0,0 106.5 -6.25 108,5 -4.25 75th percent 123.0 129.0 13.0 132.0 10.9 133.0 4,5 135.0 7.7 125.0 8.4 131.0 6.5 Maximum 139.0 145.0 20.6 139.0 21.5 140.0 11.8 140.0 13,4 143.0 10,4 140.0 10,4

The effect of hematan on blood supply to the brain of rats after global transient ischemia.

Experiments with anesthetized rats under conditions of global transient cerebral ischemia showed that 5 mg / kg gimantan with intravenous administration causes an increase in local cerebral blood flow in the rat cerebral cortex by an average of 30% from the initial level. The effect of the drug is observed 10 minutes after administration and persists throughout the entire experiment (table 4). It should be noted that the level of blood pressure under the influence of gimantan in these experiments practically does not change (table 5).

Table 5 Effect of gimantan (10 mg / kg, intravenously) on blood pressure (BP) in rats after global transient ischemia (mmHg; percentage relative initial level,%) No. op. AD f HELL Ischemia HELL Reperfusion HELL 40 min after reperfusion. F2 HELL 10 min after heimantan HELL 20 min after heimantan HELL 30 min after heimantan HELL 40 min after heimantan HELL 50 min after heimantan HELL 60 min after heimantan HELL 70 min after heimantan HELL 80 min after heimantan HELL 90 min after heimantan cu % meas. to F1 cu % meas. to F1 cu % meas. to F1 cu % meas. to f2 cu % meas. to f2 cu % meas. to f2 cu % meas. to f2 cu % meas. to f2 cu % meas. to f2 cu % meas. to f2 cu % meas. to f2 cu % meas. to f2 Min 77 thirty -72.0 90 3,1 77 -20.6 80 -19.3 81 -11.8 80 -12.7 78 -7.4 75 -8.3 71 -21.6 66 -24.1 61 -30.2 55 -34.5 25% 80 35 -64.0 one hundred 8.1 80 -0.9 88 -8.3 90 -2.8 91 0,0 99 -2.5 98 -5.2 91 -7.4 88 -9.3 81 -3.9 76 -9.1 Honey. 97 40 * -61.1 120 * 26.2 96 4.9 91 0.9 99 1.3 one hundred 5.5 one hundred 4.6 one hundred -0.9 99 2,8 98 2,8 98 -0.9 96 -1.9 75% 108 40 -52.9 120 34,4 108 11.1 98 3.9 106 11.7 116 13.8 114 16.9 110 14.3 103 16.9 109 13.5 105 9,4 106 3.9 Max 116 45 -33.8 130 41.2 116 20,0 117 15.6 117 23.3 117 28.9 116 26.7 117 thirty 119 32,2 121 34,4 120 33.3 120 33.3 F1 - the value of LMK to ischemia, F2 - the value of LMK 30 minutes after ischemia, Min - minimum 25% - 25th percent, Honey. - median 75% - 75th percent, Max - maximum; * - p <0.05 - in comparison with background 1

In an experiment on intact anesthetized rats, an electrocardiogram and respiratory rate of animals were recorded in parallel with the registration of cerebral blood flow. It was shown that gimantan does not affect the heart rate and respiratory rate of experimental animals (tables 6, 7).

Table 6 The effect of gimantan (5 mg / kg) on heart rate in intact anesthetized rats (bpm; relative to baseline,%) No. Background 10 min after the drug 20 min after the drug 30 min after preparation 40 min after preparation 50 min after preparation 60 min after preparation bpm % bpm % bpm % bpm % bpm % bpm % Minimum 368 372.0 -10.7 372.0 -7.9 380.0 -4.9 380.0 -5.0 384.0 -5.8 380.0 -6.8 25th percent 384 376.0 -3.1 384.0 -1.0 388.0 -3.0 384.0 -0.5 388.0 0.5 392.0 -1.0 Median 402 394.0 0.05 399.0 1,0 392.0 0.45 398.0 1,0 408.0 2,4 396.0 2.15 75th percent 412 408.0 2.0 428.0 3.9 412.0 3,1 428.0 3.2 432.0 43 436.0 33 Maximum 432 420.0 5,4 464.0 12.6 448.0 5,4 448.0 8.7 456.0 10.7 440.0 6.8

Table 7 The effect of gimantan (5 mg / kg) on respiratory rate (BH) in intact anesthetized rats (breath / min) No. Background 10 min after the drug 20 min after the drug 30 min after preparation 40 min after preparation 50 min after preparation 60 min after preparation breath / min % breath / min % breath / min % breath / min % breath / min % breath / min % Minimum 68 64.0 -12.0 72.0 -8.0 72.0 -10.0 72.0 -12.0 68.0 -16.0 68.0 -10.0 25th percent 76 76.0 -5.9 76.0 -5.0 72.0 -5.9 76.0 -4.8 72.0 0,0 72.0 -4.8 Median 80 80.0 -2.5 82.0 2,5 82.0 0,0 84.0 2,8 82.0 2.65 84.0 0,0 75th percent 84 88.0 4.8 92.0 5.3 92.0 5.9 108,0 15.8 84.0 10.5 92.0 15.8 Maximum 136 120.0 35.0 128.0 35.0 128.0 35.0 136.0 47.4 152.0 40.0 164.0 40,0

Description of the drawing

Figure 1 - latent period of entry into the dark chamber in the passive avoidance reaction test when playing a commemorative trace.

The abscissa axis shows the names of animal groups: “LO” - falsely operated rats, “IG” - rats with an intracerebral hematoma, “Gym 5 iv” - animals with an intracerebral hematoma, which were administered intravenously hematan (injection form) at a dose of 5 mg / kg, “Gim 10 v / b” - animals with intracerebral hematoma, which were administered intraperitoneally with hematan (substance) at a dose of 10 mg / kg.

The ordinate axis indicates the latent period of entry into the dark chamber, seconds.

* - p <0.05; ** - p <0.01 compared with falsely operated animals;

# - p <0.05; ## - p <0.01 compared with animals with intracerebral hematoma, Student's t-test for independent samples.

Claims (1)

The use of N- (2-adamantyl) -hexamethyleneimine hydrochloride (gimantan) as a means for the treatment of cerebrovascular accidents and brain injuries.

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