RU2744758C2 - Dibenzofuran oxime derivants with anticonvulsant and neuroprotective activity - Google Patents

Abstract

FIELD: chemistry.SUBSTANCE: invention relates to a group of derivatives of O-oxime 3,4,6,7,8,9-hexahydrodibenzo[b,d]furan-1(2H)-one of a general formula, wherein.Invention also relates to use of derivatives of O-oxime 3,4,6,7,8,9-hexahydrodibenzo[b,d]furan-1(2H)-one of the abovementioned general formula, wherein,and to method of synthesis of such compounds.EFFECT: synthesis of such compounds that may be used in medicine as anticonvulsant and neuroprotective products and products with combined anticonvulsant and neuroprotective activity for treatment of epilepsy and paroxysmal conditions.5 cl, 9 tbl, 8 ex

Description

The field to which the invention relates

The invention relates to medicine, in particular to pharmacology and pharmacy, to biology and chemistry, specifically to a new group of derivatives of O-oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -on, general formula (1)

Where

which can be used as pharmacological agents for the treatment of epilepsy, paroxysmal conditions, including post-stroke epilepsy, as well as the treatment of post-stroke neurological complications.

Background of the invention

Modern antiepileptic drugs (AEDs) include substances of various classes of chemical compounds and have a different mechanism of action, many of which have been known for a long time, but none of these drugs is unique, used for all types of epilepsy. The classification of epileptic symptoms is diverse and includes a dichotomy both in the duration of the seizures and in the form: about 40 forms of epilepsy are distinguished. In Russia, today, more than 30 AEDs have been registered and approved for use, and none of them is universal, equally successfully used in any form of epileptic seizures. Despite the emergence of new AEDs, the problem of the curability of epilepsy does not lose its relevance, since the number of patients with drug-resistant epilepsy remains significant, accounting, according to various estimates, 25-30% of the total number of people with this disease (Karlov V.A., 2010a; Kwan P., Brodie M. J., 2000). Treatment of epilepsy is carried out continuously for many years, and sometimes throughout the patient's life, while AEDs have various side effects (Karlov V.A.Epilepsy in children and adult women and men: a guide for doctors / V.A. : Medicine, 2010 .-- 717 s).

One of the most common risk factors for developing symptomatic epilepsy is cerebrovascular disease. A decrease in the seizure threshold after a stroke is one of the manifestations of neurological deficit. According to the International Antiepileptic League, the incidence of symptomatic focal vascular epilepsy is 6-8%. The development of early epileptic seizures has a negative impact on the recovery period of stroke, predisposes to the persistence of the severity of neurological deficit, low survival rates and the risk of recurrent stroke within two years after the primary stroke. Patients with cerebral ischemia often develop focal seizures, and in the first 7 days of stroke, simple partial seizures prevail. Early epileptic seizures are more often observed with the cardioembolic subtype of stroke and with damage to the left carotid basin.

Benzofuran derivatives have a wide range of pharmacological effects and are well known in Russia and abroad. In the Research Institute of Pharmacology of the Russian Academy of Medical Sciences were synthesized and studied derivatives of 5-nitrobenzofuran with neurotropic action of the deprimating type, including with anticonvulsant activity in one of the compounds [Patent 770040. Derivatives of benzofuran with neurotropic action of the deprimating type // 1979]. Derivatives of dibenzofuranol, obtained at the Institute of Pharmacology, have adrenergic blocking, hypotensive, antispasmodic, neurotropic-depriving activity [Ed. Certificate No. 869278, 1980]. Also at the Institute of Pharmacology were obtained derivatives of dibenzofuranol with antiarrhythmic and antianginal activity [Ed. Certificate No. 1132507, 1983]. Among benzofurans known drug cordaron, which has interferon inductive activity, as well as other derivatives with immunomodulatory and neuroprotective activity [Application WO2006129318 A3. EP 1885709 A2. Benzofuran derivatives with therapeutic activities // 2006].

M,

J, Silva-Grecchi T. et al. A Natural Benzofuran from the Patagonic Aleurodiscus vitellinus Fungus has Potent Neuroprotective Properties on a Cellular Model of Amyloid-β Peptide Toxicity // J Alzheimers Dis. 2018;61(4):1463-1475] и противоопухолевой активностью в отношении клеток Меланомы [Zhu JJ. Yan GR1, Xu ZJ, Hu X et al. Inhibitory Effects of (2'R)-2',3'-dihydro-2'-(1-hydroxy-1-methylethyl)-2,6'-bibenzofuran-6,4'-diol on Mushroom Tyrosinase and Melanogenesis in B16-F10 Melanoma Cells // Phytother Res. 2015 Jul; 29(7):1040-5]. Среди дибензофуранов синтезированы структурные аналоги лигандов каннабинойдных рецепторов, демонстрирующих к ним сродство [Sadia Akram. Synthesis of Dibenzofurans via a Palladium Catalyzed Oxidative Ring Closure Reaction // May 2013; Заявка EP 1885709 A2. Benzofuran derivatives with therapeutic activities // 2006]. Наиболее известное производное дибензофуранов (usnic acid, 2,6-diacety 1-7,9-dihydroxy-8,9bdimethyl-1,3 [2H,9bH]-dibenzofurandione), выделенное из лишайников, обладает рядом фармакологических эффектов, таких как, гастропротекторным [Odabasoglu, F.; Cakir, A.; Suleyman, Н. Asian, A.; et al. Gastroprotective and antioxidant effects of usnic acid on indomethacin-induced gastric ulcer in rats. J. Ethnopharmacol. 2006, 103, 59-65. Molecules 2014, 19 14525], кардиологическим [Behera, B.C.; Mahadik, N., Morey, M. Antioxidative and cardiovascular-protective activities of metabolite usnic acid and psoromic acid produced by lichen species Usnea complanata under submerged fermentation. Pharm. Biol. 2012, 50, 968-979], цитопротекторным [De Paz, G.A.; Raggio, J.;

, M.P.; et al. HPLC isolation of antioxidant constituents from Xanthoparmelia spp. J. Pharm. Biomed. Anal. 2010, 53, 165-171], иммуностимулирующим [Santos, L.C.; Honda, N.K.; Carlos, I.Z.; Vilegas, W. Intermediate reactive oxygen and nitrogen from macrophages induced by Brazilian lichens. Fitoterapia 2004, 75, 473-479], противомикробным [

, В.;

, M;

, Т.; et al. Biological Activities of Toninia candida and Usnea barbata Together with Their Norstictic Acid and Usnic Acid Constituents. Int. J. Mol. Sci. 2012, 13, 14707-14722], противовоспалительным [Jin, J.Q.; Li, C.Q.; He, L.C. Down-regulatory effect of usnic acid on nuclear factor-kappaB-dependent tumor necrosis factor-alpha and inducible nitric oxide synthase expression in lipopolysaccharide-stimulated macrophages RAW 264.7. Phytother. Res. 2008, 22, 1605-1609] и антиканцерогенным [Brisdelli, F.; Perilli, M.; Sellitri, et al. Cytotoxic activity and antioxidant capacity of purified lichen metabolites: An in vitro study. Phytother. Res. 2013, 27, 431-437;

, M.;

, M.;

, J.; et al. Variable responses of different human cancer cells to the lichen compounds parietin, atranorin, usnic acid and gyrophoric acid. Toxicol. In Vitro 2011, 25, 37-44] действием, в основном благодаря антиоксидантной активности в восстановлении окислительного стресса. Синтезированны дибензофураны, обладающие противовоспалительной активностью [Заявка CN 102887880 A1. Dibenzofuran derivatives, and preparation method and application thereof//2012].Currently, benzofuran derivatives ((2'R) -2 ', 3'-dihydro-2' - (1-hydroxy-1-methylethyl) -2,6'-bibenzofuran-6,4'-diol (1) and 5,6-dimethoxy-2- (3-hydroxy-5-methoxyphenyl) benzofuran) from Patagonic Aleurodiscus vitellinus Fungus and Morus notabilis, which have neuroprotective activity against β-amyloid toxicity [Hu X, Wang M, Yan GR et al. 2-Arylbenzofuran and tyrosinase inhibitory constituents of Morus notabilis // J Asian Nat Prod Res. 2012; 14 (12): 1103-8;

M,

J, Silva-Grecchi T. et al. A Natural Benzofuran from the Patagonic Aleurodiscus vitellinus Fungus has Potent Neuroprotective Properties on a Cellular Model of Amyloid-β Peptide Toxicity // J Alzheimers Dis. 2018; 61 (4): 1463-1475] and anti-tumor activity against Melanoma cells [Zhu JJ. Yan GR1, Xu ZJ, Hu X et al. Inhibitory Effects of (2'R) -2 ', 3'-dihydro-2' - (1-hydroxy-1-methylethyl) -2,6'-bibenzofuran-6,4'-diol on Mushroom Tyrosinase and Melanogenesis in B16 -F10 Melanoma Cells // Phytother Res. 2015 Jul; 29 (7): 1040-5]. Among dibenzofurans, structural analogs of cannabinoid receptor ligands have been synthesized, demonstrating an affinity for them [Sadia Akram. Synthesis of Dibenzofurans via a Palladium Catalyzed Oxidative Ring Closure Reaction // May 2013; EP 1885709 A2. Benzofuran derivatives with therapeutic activities // 2006]. The most famous derivative of dibenzofurans (usnic acid, 2,6-diacety 1-7,9-dihydroxy-8,9bdimethyl-1,3 [2H, 9bH] -dibenzofurandione), isolated from lichens, has a number of pharmacological effects, such as gastroprotective [Odabasoglu, F .; Cakir, A .; Suleyman, H. Asian, A .; et al. Gastroprotective and antioxidant effects of usnic acid on indomethacin-induced gastric ulcer in rats. J. Ethnopharmacol. 2006, 103, 59-65. Molecules 2014, 19 14525], cardiac [Behera, BC; Mahadik, N., Morey, M. Antioxidative and cardiovascular-protective activities of metabolite usnic acid and psoromic acid produced by lichen species Usnea complanata under submerged fermentation. Pharm. Biol. 2012, 50, 968-979], cytoprotective [De Paz, GA; Raggio, J .;

, MP; et al. HPLC isolation of antioxidant constituents from Xanthoparmelia spp. J. Pharm. Biomed. Anal. 2010, 53, 165-171], immunostimulatory [Santos, LC; Honda, NK; Carlos, IZ; Vilegas, W. Intermediate reactive oxygen and nitrogen from macrophages induced by Brazilian lichens. Fitoterapia 2004, 75, 473-479], antimicrobial [

, IN.;

, M;

, T .; et al. Biological Activities of Toninia candida and Usnea barbata Together with Their Norstictic Acid and Usnic Acid Constituents. Int. J. Mol. Sci. 2012, 13, 14707-14722], anti-inflammatory [Jin, JQ; Li, CQ; He, LC Down-regulatory effect of usnic acid on nuclear factor-kappaB-dependent tumor necrosis factor-alpha and inducible nitric oxide synthase expression in lipopolysaccharide-stimulated macrophages RAW 264.7. Phytother. Res. 2008, 22, 1605-1609] and anticarcinogenic [Brisdelli, F .; Perilli, M .; Sellitri, et al. Cytotoxic activity and antioxidant capacity of purified lichen metabolites: An in vitro study. Phytother. Res. 2013, 27, 431-437;

, M .;

, M .;

, J .; et al. Variable responses of different human cancer cells to the lichen compounds parietin, atranorin, usnic acid and gyrophoric acid. Toxicol. In Vitro 2011, 25, 37-44] action, mainly due to antioxidant activity in the restoration of oxidative stress. Synthesized dibenzofurans with anti-inflammatory activity [Application CN 102887880 A1. Dibenzofuran derivatives, and preparation method and application thereof // 2012].

Technical result

The present invention is based on the problem of creating compounds - derivatives of dibenzofuranone oxime with high anticonvulsant activity and neuroprotective activity.

Compounds of general formula (1), their properties and preparation method are not described in the special and patent literature.

The proposed technical solution - derivatives of O- (R) -oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -one (1):

O- (3,4-dichlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1а)

O- (4-cinnamoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1b)

O- (4-chlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1c)

O- (4-fluorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1d)

O- (2-chlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1e)

O- (3-chlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1е)

O- (2,4-dichlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1g)

O- (4-chlorophenoxyacetyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -one (1h).

Obtaining the claimed compounds

Compounds of the general formula can be obtained according to the following general scheme:

The reaction of 2-chlorocyclohexanone (2) with 1,3-cyclohexanedione (3) in methanol followed by treatment with 10% NaOH, extraction with chloroform and distillation gives 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan -1 (2H) -one (4) (DBP), which is converted to oxime BDF (5) by its reaction with hydroxylamine hydrochloric acid in the presence of NaOH. Boiling for 3-5 hours oxime (5) in benzene with acid chlorides of the corresponding acids in the presence of triethylamine gives O- (R) -oximes 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -one (1a-h).

Compounds 1 are white crystalline substances, insoluble in water, soluble in some organic solvents such as DMSO. The structure of substances of general formula 1 is confirmed by the data of NMR spectra, and their purity by the data of elemental analysis.

O-oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -one (5) [Application WO 2017/109724 A1]

To a solution of 9.00 g (0.047 mol) of dibenzofuranone (4) [Application WO 2017/109724 A1] in 34.4 ml of alcohol add 5.17 g (0.070 mol) of hydroxylamine hydrochloric acid in 3.5 ml of water (precipitation occurs), while stirring, solid NaOH (crushed) is added in small portions. Bring to a boil, boil for 5 minutes, cool and pour a mixture of 94.7 ml of 10% hydrochloric acid while cooling with water, filter off the precipitate 8.75 g (97%) (5) melting point 192-194 ° C (alcohol). Rf = 0.66 [9.5: 0.5 chlf: cn].

Calculated%: C, 70.21; H, 7.37; N, 6.82. C ₁₂ H ₅ NO ₂

Found%: C, 70.21; H, 7.37; N, 7.04.

PMR spectrum (DMSO-d ₆ ) δ cm ^-1 : 1.55-1.90, 2.40-2.70 (14H, m-ring)

Example 1

O- (3,4-dichlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1а)

To 1.53 g (7.5 mmol) of BDF (5) oxime in 10 mg of dry benzene, 1.72 g (8.2 mmol) of 3,4-dichlorobenzoic acid chloride are added in the presence of 0.83 g (8.2 mmol ) dry triethylamine is boiled for 4 hours, the precipitate is filtered off (ch.g. Et ₃ N-soluble in water); the benzene solution is evaporated, the residue is treated with a NaHCO ₃ solution, 2.50 g is filtered off, and crystallized from alcohol. 2.10 g (88%) are obtained, melting point 158-159 ° C. Rf = 0.90 [9.5: 0.5 chlf: cn].

Calculated%: C 60.33; H 4.53; Cl 18.75; N 3.70. C ₁₉ H ₁₇ Cl ₂ NO ₃

Found%: C 60.59; H 4.66; Cl 18.65; N 3.66.

PMR spectrum (DMSO-d ₆ ) δ cm ^-1 : 1.67-1.68 (4-H, two m CH ₂ - 7.8), 1.76-1.94 (2H, quintet J = 6, 2, CH ₂ - 3), 2.50-2.53 (4H, two t, J = 6.2, CH ₂ - 6.9), 2.71 (2H, t, J = 6.2, CH ₂ - 1), 2.86 (2H, t, J = 6.2, CH ₂ - 2), 7.82-7.86 (1H, d, J = 7.85, CH-5), 7, 95-7.97 (1H, d, J = 8.5, CH-6), 8.14 (1H, s, CH-2).

Example 2

O- (4-cinnamoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (16)

It was obtained similarly to 1а from 1.03 g of DBP oxime (5) and cinnamic acid chloride. Yield 1.40 g (84%). Melting point 137-138 ° C, Rf = 0.80 [9.5: 0.5 chlf: cn].

Calculated%: C, 75.19; H, 6.31; N, 4.12. C ₂₁ H ₂₁ NO ₃

Found%: C, 75.36; H, 6.48; N, 4.07.

PMR spectrum (DMSO-d ₆ ) δ cm ^-1 : 1.58-2.83 (14-H, m, (CH ₂ ) ₃ , (CH ₂ ) ₄ and rings), 6.79-7.15 ( 2H, two d, J = 15.8, CH = CH), 7.45 and 7.15 (5H, m, ArH).

Example 3

O- (4-chlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1c)

It was obtained similarly to 1а from 1.40 g of DBP oxime (5) and 4-chlorobenzoic acid chloride. Yield 1.80 g (77%), melting point 150-151 ° C (from alcohol). Rf = 0.85 [9.5: 0.5 chlf: cn].

Calculated%: C 66.37; H 5.28; C1 10.31; N 4.07. C ₁₉ H ₁₈ ClNO ₃

Found%: C 66.2; H 5.31; Cl 10.15; N 3.99.

PMR spectrum (DMSO-d ₆ ) δ cm ^-1 : 1.68 and 1.76 (4-H, two m, CH ₂ - 7.8), 1.94 (2H, m, CH ₂ - 3), 2.54 (4H, m, CH ₂ - 6.9), 2.71 and 2.84 (4H, m, CH ₂ - 2.4), 7.49 (2H, d, J = 8.5, CH-3 ', 5'), 8.05 (4H, d, J = 8.5 CH-2 ', 6')

Example 4

O- (4-fluorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1d)

It was obtained similarly to 1а from 1.53 g of DBP oxime (5) and 4-fluorobenzoic acid chloride. Yield 2.38 g (nec.), Crystallized from alcohol. Yield 1.90 g (80%). Melting point 119-120 ° C (from alcohol). Rf = 0.85 [9.5: 0.5 chlf: cn]. Rf = 0.6 (hlf)

Calculated%: C 69.71; H 5.54; F 5.80; N 4.28. C ₁₉ H ₁₈ FNO ₃

Found%: C 69.71; H 5.54; F 5.80; N 4.28.

PMR spectrum (DMSO-d ₆ ) δ cm ^-1 : 1.68 and 1.69 (4-H, two m, CH ₂ - 7.8), 1.94 (2H, quintet, J = 6.4, CH ₂ - 3), 2.51 (4H, m, CH ₂ - 6.9), 2.71 (2H, t, J = 6.4, CH ₂ - 2), 2.85 (2H, t, J = 6.4, CH ₂ - 4), 7.39 (2H, m, CH-3 ', 5'), 8.08 (2H, m, CH-2 ', 6')

Example 5

O- (2-chlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1e)

It was obtained similarly to 1а from 1.03 g of DBP oxime (5) and 2-chlorobenzoic acid chloride. Yield 1.40 g (82%), crystallized from alcohol. Melting point 123-125 ° C (from alcohol). Rf = 0.85 [9.5: 0.5 chlf: cn]. Rf = 0.6 (hlf)

Calculated%: C 66.37; H 5.28; C1 10.31; N 4.07. C ₁₉ H ₁₈ ClNO ₃

Found%: C 66.24; H 5.33; C1 10.02; N 4.09.

PMR spectrum (CDCl ₃ ) δ cm ^-1 : 1.75 and 1.76 (4H, two m, CH ₂ - 7.8), 1.82-1.83 (2H, m, CH ₂ -3), 2.58 and 2.66 (4H, m, CH ₂ - 6.9), 2.74 and 2.88 (4H, two m, CH ₂ - 2.4), 7.39 and 7.41 (3H , two m, CH- 3 ', 4', 5 '), 7.89 (1H, m, CH-6')

Example 6

O- (3-chlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1е)

It was obtained similarly to 1а from 1.75 g of DBP oxime (5) and 3-chlorobenzoic acid chloride. Yield 0.85 g (68%), crystallized from alcohol. Melting point 168-170 ° C. Rf = 0.8 (chlf)

PMR spectrum (CDCb) δ cm ^-1 : 1.75 and 1.76 (4H, two m, CH ₂ - 7.8), 1.84 (2H, m, CH ₂ - 3), 2.58 (4H , t, CH ₂ - 6), 2.66 and 2.75 (4H, two m, CH ₂ - 4.9), 2.88 (2H, m, CH ₂ - 2), 7.59 and 7, 60 (2H, two m, CH-4 ', 5'), 7.99 (2H, m, CH-2 ', 6').

Example 7

O- (2,4-dichlorobenzoyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1g)

It was obtained similarly to 1а from 1.03 g of DBP oxime (5) and 2,4-dichlorobenzoic acid chloride. Yield 1.30 g (68%). Melting point 149-150 ° C.

Calculated%: C 57.58; H 4.83; Cl 17.89; N 3.53. C ₁₉ H ₁₇ NO ₃ Cl ₂ ⋅H ₂ O

Found%: C 57.76; H 4.84; Cl 17.88; N 3.73

PMR spectrum (CDCb) δ cm ^-1 : 1.60-1.76 (4H, two m, CH ₂ - 7.8), 176-1.85 (2H, m, CH ₂ - 3), 2.01 -2.03 (2H, m, CH ₂ - 9), 2.58-2.7 (4H, m, CH ₂ - 2.6), 2.84 (2H, m, CH ₂ -4), 7 , 34 (1H, m, CH-3 '), 7.51 (1H, s, CH-5'), 7.81-7.84 (1H, d, J = 6.4, CH-2 ')

Example 8

O- (4-chlorophenoxyacetyl) oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2Н) -one (1h)

It was obtained similarly to 1а from 1.03 g of DBP oxime (5) and 2- (4-chlorophenoxy) acetic acid chloride. Yield 1.60 g (86%). Melting point 117-118 ° C.

Calculated%: C 64.25; H 5.39; Cl 9.48; N 3.74. C ₂₀ H ₂₀ ClNO ₄

Found%: C 64.33; H 5.44; Cl 9.44; N 3.65.

PMR spectrum (DMSO) δ cm ^-1 : 1.60 (4-H, m, CH ₂ - 7.8), 1.98 (2H, m, CH ₂ - 3), 2.52 (4H, m, CH ₂ - 6.9), 2.70-2.80 (4H, m, CH ₂ - 2.4), 5.02 (2H, s, OCH ₂ ), 7.00-7.46 (5H, m, ArH).

Experimental pharmacological part

The organization and implementation of the work was carried out in accordance with the order of the Ministry of Health of Russia No. 199 dated April 01, 2016 "On the approval of the rules of good laboratory practice." The animals were kept in accordance with SP 2.2.1.3218-14 "Sanitary and Epidemiological Requirements for the Arrangement, Equipment and Maintenance of Experimental Biological Clinics (Vivariums)" dated August 29, 2014 No. 51. The experiments were approved by the Commission on Biomedical Ethics of the V.V. Zakusov "(Minutes No. 6 dated April 16, 2018).

The results were statistically processed using MS Excel 2010 and BioStat 2009 (Analyst Soft Inc.). Normality of data distribution was determined using the Shapiro-Wilk test. The significance of differences in values between groups was determined using nonparametric criteria: Kruskal-Wallis and Fisher's exact test.

Example 1

Anticonvulsant action of dibenzofuranone oxime derivatives in a model of primary generalized seizures caused by maximal electroshock (MES)

The experiments were carried out on white outbred male mice, weighing 20-26 g, obtained from the Stolbovaya nursery, State Institution NTsBMT (Moscow Region). Each dose of the compound was tested in 6-25 animals.

The anticonvulsant effect of furan derivatives was investigated on models of primary generalized seizures caused by maximum electroshock (MES), simulating the so-called "large" (Grantmal) seizures (Voronina TA., Nerobkova LN Guidelines for studying the anticonvulsant activity of pharmacological substances. " Guidelines for conducting preclinical studies of drugs "Part. 1. FSBI" NTsEMSP ". Moscow, Publishing house GrifiK, 2012, Chapter 14, pp. 235-250; Loscher et al., The role of technical, biological and pharmacological factors in the laboratory evaluation of anticonvulsant drugs. II. Maximal electroshock seizure models, Epilepsy Res., 1991, v. 8, p. 79-94; Swinyard EA - Laboratory evaluation of antiepileptic drugs. Review of laboratory methods, Epilepsia, 1969, v. 10, pp. 107-119; Loscher W., 1991; Rostock A., 1996).

An electroconvulsive seizure (ES) was created using a certified Rodent ShockerRS device, type 221 (HarvardApparatus, GmbH, Germany). Animals received electrical stimuli through special corneal electrodes (mode 250 V / mA: 12/13/14 mA, duration 0.2 s). The following parameters were recorded: tonic extension of the hind and forelimbs and death of the animals. The anticonvulsant effect of the claimed compounds was assessed by the ability to prevent the development of tonic extension and death of animals. The test compounds were dissolved in saline with Tween80 and injected intraperitoneally (ip) 40 minutes before the MES. The reference drug Mexidol (substance) was dissolved in saline and administered at doses of 100 and 200 mg / kg in a similar regimen. The effectiveness of the compounds was assessed by the increase in the number of animals without tonic seizures and the proportion of survivors (%) after MES relative to the corresponding control groups.

Compound 1a was investigated at doses of 5-100 mg / kg, comparing the obtained values with those of the control group No. 1 (table 2). It was found that compound 1а with a single dose of 20 mg / kg significantly increased the survival rate by 36%, and at doses of 30, 40, 60 and 100 mg / kg - by 50% relative to the corresponding control with MES (control No. 1). At doses of 5, 10 and 80 mg / kg, compound 1a had no effect on the survival rate (Table 1). Compound 1b, when administered to mice at a dose of 20 and / kg, contributed to an increase in the survival rate of animals by 35% (P≤0.1) relative to control No. 2, however, when the dose was increased by 2 times, the effectiveness fell (table 1). Compound 1c at doses of 30 and 40 mg / kg contributed to the elimination of tonic seizures and an increase in the survival rate of animals by 45.6-66.5% compared to the control group No. 3. With increasing dose (60 mg / kg), the efficacy of the compound slightly decreased, while the protection against death was 43%, while in the low dose range (5 and 20 mg / kg), no efficacy was observed (Table 1). Compound 1e at a dose of 40 mg / kg reduced the number of animals with tonic-clonic seizures caused by MES, increasing the survival rate of mice to 72% (in control No. 3 - 21%), however, it did not interfere with the action of MES when the dose was reduced to 20 mg / kg (Table 1). Compound 1e at a dose of 20 mg / kg contributed to an increase in the survival rate of animals after MES by 40% (P≤0.1) compared to the control group No. 4, while when the dose was increased to 40 mg / kg, no protective effect against death was observed (Table 1) ... Compound 1h at a dose of 50 mg / kg contributed to an increase in the survival rate of animals by 50% (P≤0.1) relative to the values of the control group No. 5, while at doses of 5 and 20 mg / kg no protective effect was observed in the development of convulsive manifestations and, as a consequence, the survival rate did not increase (table 1). An increase in the survival rate of mice by 42% (P≤0.05) after seizures caused by MES was observed with the introduction of compound 1g at a dose of 60 mg / kg. With a decrease in the dose to 40 mg / kg, the effectiveness of compound 1g fell, which was manifested in a decrease in the survival rate of animals to 39% (p≤0.1) relative to control values No. 6 (table 1). The administration of compound 1g at doses of 2.5, 20 and 40 mg / kg did not develop the anticonvulsant effect necessary to stop the seizure reactions caused by the MES relative to the control values No. 7 (Table 1). Mexidol at a dose of 100 mg / kg did not affect the severity of convulsive manifestations and did not prevent the death of animals. With the introduction of Mexidol at a dose of 200 mg / kg, the death of animals was statistically significantly reduced by 62.5% compared to control No. 8 (Table 1).

* - reliability of differences from the control group, at P≤0.05 (Fisher's exact test); # - the tendency towards the reliability of differences from the control group, at P≤0.1 (Fisher's exact test).

Thus, it was found that for compound 1a the range of effective doses is wide and corresponds to 20-100 mg / kg. The survival rate of animals with the introduction of compound 1a increased by a maximum of 50% relative to the control values. Compounds 1e (40 mg / kg) and 1c (30 mg / kg) demonstrated statistically significant anticonvulsant activity, the administration of which increased the survival rate of mice by 51% and 66.5%, respectively. Compounds 1f (20 mg / kg), 1h (50 mg / kg), and 1b (20 mg / kg) showed tendency level activity in the MES antagonism test. Comparison drug Mexidol increases survival by 62.5% when administered at a dose of 200 mg / kg.

Example 2

Anticonvulsant action of dibenzofuranone oxime derivatives in a model of primary generalized seizures caused by corazole

The experiments were carried out on white outbred male mice weighing 20-26 g. Each dose of the compound was studied in 8-12 animals. Antagonism test with corazole (pentylenetetrazole, Sigma-Aldrich, USA) - a GABA receptor antagonist is the basic method for assessing the action of substances with anticonvulsant activity (Voronina T.A., Nerobkova L.N. Guidelines for studying the anticonvulsant activity of pharmacological substances. on conducting preclinical studies of drugs "Part. 1. FSBI" NTsEMSP ". Moscow, Publishing house GrifiK, 2012, Chapter 14, pp. 235-250; Loscher et al., The role of technical, biological and pharmacological factors in the laboratory evaluation of anticonvulsant drugs. III. Pentylenetetrazolseizuremodels. Epilepsy Res., 1991, v. 8, pp. 171-189). In this technique, seizures are caused by chemical action and simulate primary generalized seizures in the so-called "small" (Petit mal) seizures. Experimental groups of animals were injected intravenously with the test substances dissolved in physiological solution with Tween 80 40 minutes before corazole. The reference drug Mexidol (substance) was dissolved in saline and administered at doses of 100 and 200 mg / kg in a similar regimen. To create corazole seizures, animals were injected subcutaneously into the cervical spine with corazole (pentylenetetrazole) at a dose of 95 mg / kg. The development of seizures and death of animals was recorded for 120 min. The efficacy of the compounds was assessed by the increase in the proportion of surviving mice after corazole relative to the corresponding control groups.

It was found that in the groups of control animals (No. 1, 2 and 3), after administration of corazole at a dose of 95 mg / kg, convulsive manifestations developed in the following sequence: one or more myoclonic twitchings of the whole body in 100% of mice; repetitive clonic seizures of the fore and / or hind limbs lasting more than 3 seconds without loss of the flipping reflex in 100% of mice; generalized clonic convulsions of the fore and hind limbs with a loss of the rolling reflex in 90% of mice; death of animals - 55-90% of mice.

Compound 1a at doses of 5, 20, and 40 mg / kg had no effect on corazole-induced convulsions and the survival rate (Table 2). Compound 1 at doses of 20 and 40 mg / kg with a single administration had no effect on the number of surviving animals (Table 2). When compound 1e was administered at a dose of 20 mg / kg, 100% survival of animals was observed relative to the corresponding values of the control group No. 2 (P <0.05). When this compound was administered at a dose of 40 mg / kg, there were no significant changes in convulsive reactions relative to the control values (Table 2). Compound 1g at a dose of 40 mg / kg showed weak anticonvulsant activity relative to control No. 2 in the test of corazole seizures, slightly increasing the survival rate of animals by 22.2% (table 2). With the introduction of compounds 1c (10, 20, 30 mg / kg) and 1g (10, 20, 40 mg / kg), as well as 1h at doses of 20 and 40 mg / kg, no protective effect was observed against the development of convulsive reactions and death caused by corazole, when compared with the control group No. 3 (table 2). Mexidol at a dose of 100 mg / kg did not change the survival rate of animals, but at a dose of 200 mg / kg increased the proportion of surviving animals by 42% relative to the control values of group No. 4 (table 2).

& - reliability of differences from the control group, at Р≤0.05 (Fisher's exact test); # -the tendency to the reliability of differences from the control group, at P≤0.1 (Fisher's exact test).

Thus, it was found that, in the test of antagonism with corazole, compound 1e at a dose of 20 mg / kg protects 100% against animal death and the development of generalized convulsive manifestations. Compounds 1a, 1b, 1c, 1g, and 1h in the studied doses did not show significant changes in the survival rate of animals in the test of antagonism with corazole. Compound 1 g at a dose of 40 mg / kg exhibits weak anticonvulsant activity, increasing the survival rate of mice by 22.2%. The reference drug Mexidol (200 mg / kg) with a tendency towards statistical significance increases the number of surviving animals by 42%.

Example 3

Antihypoxic effect of dibenzofuranone oxime derivatives on the model of hypoxia with hypercapnia in a pressurized volume, model of canned hypoxia

The experiments were carried out on white outbred male mice weighing 22-26 g. Each dose of the compound was studied in 8-10 animals.

The antihypoxic effect of furan derivatives was studied using a model of hypoxia with hypercapnia in a hermetic volume, a model of canned hypoxia. The animals were placed in a 250 ml jar, tightly closed with a lid, hermetically fixed with a paraffin film. The life expectancy of animals was recorded until the last atonal breath using a stopwatch (Voronina T.A., 2012: Roshchina L.F., Ostrovskaya R.U. 1981). Experimental groups of animals were injected intravenously with the test substances dissolved in physiological solution with Tween 80 40 minutes before the test. The reference drug Mexidol (substance) was dissolved in saline and administered at doses of 100 and 200 mg / kg in a similar regimen. The effectiveness of the compounds was assessed by the increase in the animal life time relative to the corresponding control groups.

When studying the antihypoxic effects of dibenzofuran derivatives in a model of normobaric hypoxia with hypercapnia, it was found that compound 1а statistically significantly increased the life span of animals relative to the control values of group No. 1: by 3.2 minutes at a dose of 10 mg / kg and 7.4 minutes at a dose of 30 mg / kg (table 3). Compounds 1c (at a dose of 30 mg / kg), 1 g (at doses of 20, 40, and 60 mg / kg), 1b (at doses of 20, 40, and 60 mg / kg) and 1e (at doses of 20 and 40 mg / kg) did not cause an increase in the lifespan of mice in comparison with the corresponding control groups (No. 2, 3, 4) under conditions of normobaric hypoxia with hypercapnia in the hermetic volume (Table 3). Mexidol at a dose of 100 mg / kg did not change the survival rates of animals relative to the control values of group No. 5, but statistically significantly increased the lifespan of mice (by 2.7 minutes) when the drug was administered at a dose of 200 mg / kg (Table 3).

* - reliability of differences from the control group, at P≤0.05 (Kruskal-Wallis test)

It was found that in the test of hypoxia with hypercapnia in the hermetic volume, compound 1a possesses statistically significant antihypoxic activity. Its effectiveness increases with increasing dose from 10 to 30 mg / kg, contributing to an increase in the life time of animals by 3.2 and 7.4 minutes. Mexidol at a dose of 200 mg / kg increases the life span of animals by 2.7 minutes.

Example 4

Neuroprotective properties of dibenzofuranone oxime derivatives in the model of global ischemia, model of ischemic stroke.

The experiments were carried out on male rats weighing 240-260 g according to the "Guidelines for conducting preclinical studies" (Publication of the Federal State Budgetary Institution "NTsEMSP" of the Ministry of Health and Social Development of Russia, Moscow, 2012, Part 1: Chapter 28; Mirzoyan R.S. et al., 2012; Voronina T.A. et al., 2012b). Each group contained 8 to 29 animals. To simulate ischemic stroke (IS) in rats anesthetized with chloral hydrate (350 mg / kg, i.p.), one-stage bilateral ligation of the common carotid arteries was performed to the site of bifurcation (global ischemia model). For this, a skin incision was made in the rats in the neck area, followed by extraction of 2 carotid arteries and their separation from the vagus nerve. Then the carotid arteries were simultaneously tied with a silk ligature and the wound was sutured. For sham-operated animals (LO), similar surgical procedures were performed, but without ligation of the carotid arteries. Compounds 1a, 1b, 1d, 1e, 1g were administered 2-3 hours after surgery at a dose of 10 mg / kg and then once a day for 6 days. The test compounds were dissolved in physiological solution with Tween-80. The control groups with IS (No. 1, 2) and LO were injected with physiological saline with Tween-80 in an equivalent volume. The reference drug Mexidol (100 mg / kg) was dissolved in physiological saline. The control group No. 3 was injected with an equivalent volume of solvent. Survival of animals after surgery and the development of neurological deficits (Stroke-index McGrow modified by Ganushkina (Ganushkina, 1998)) were recorded for 14 days: 0- (7-8 hours after surgery), 1- (after 24 hours), 2-, 3-, 7- and 14th days after surgery.

It was found that compound 1а at a dose of 10 mg / kg with a single injection 2 hours after ischemic stroke protected animals from death for 7-8 hours after surgery (day 0), while in the control group with IS No. 1, 2 -e animals (table 4). The weakening of the neurological deficit observed during the experiment (table 5, 6), in the group of rats receiving compound 1a for the next 6 days after the operation, led to a decrease in the death of animals by 12% (3 days) and by 37% (7 and 14 day) in comparison with the control group II No. 1 (table 4). Thus, by the end of the experiment, the total death of animals with IS, which received compound 1a (10 mg / kg), was 55%, while in the control group of rats with IS No. 1 without therapy, 92% of rats died (Table 4). In the group of rats that received a single dose of compound 1b (10.0 mg / kg), no death was observed on the day of surgery (day 0), while in the control group with IS No. 2, 6 animals (21%) died. Compound 1b, after repeated administration (10 mg / kg for 6 days), with a tendency towards statistical significance, protected rats from death by 7- (by 30%) and statistically significant on the 14th day (by 33%) of recording post-stroke dynamics (Table 4) ... Compound 1b statistically significantly weakened the registered neurological disorders, which contributed to a decrease in the number of animals with circus movements and paresis of the extremities, as well as weakening of mild neurological symptoms (ptosis and limb weakness) (Table 5, 6). Compounds 1d, 1e and 1g, when administered to animals within 7 days after surgery, did not lead to a decrease in neurological deficit and a decrease in the death of animals within 14 days of recording post-stroke dynamics (Table 4, 5, 6). Mexidol at a dose of 100 mg / kg protected animals from death during the first 7 days of administration after surgery and by the 14th day increased the survival rate of rats with ischemia by 21.5% relative to the group of control animals with IS No. 3 (Table 4).

Notes: AI-ischemic stroke; LO-falsely operated. * - reliability of values in comparison with the control group with IS, at p <0.05 (Fisher's exact test); # - the trend towards the reliability of values compared to the control group with IS, at p <0.1 (Fisher's exact test).

Notes: AI-ischemic stroke; LO-falsely operated; p / ptosis - semiptosis. * - reliability of values compared with the control group, at p≤0.05 (Fisher's exact test).

Notes: AI-ischemic stroke; LO-falsely operated; p / ptosis - semiptosis * - reliability of values in comparison with the control group, at p <0.05 (Fisher's exact test); # - the trend towards the reliability of values compared to the control group with IS, at p <0.1 (Fisher's exact test).

Thus, compounds 1а (10 mg / kg / 7 days of administration) and 1b (10 mg / kg / 7 days of administration) have a pronounced neuroprotective effect on the model of ischemic stroke, in the groups of rats of which, on the 14th day after IS, the survival rate increased by 37%. (compound 1a) and 33% (compound 1b) compared with the corresponding control groups with IS. Compound 1b in this experiment was the most effective, since it not only reduces the death of animals, but also statistically reliably weakens neurological deficit. Compounds 1d, 1e, and 1g (10 mg / kg / 7 days of administration) do not protect against death of animals after modeling IS and the development of postischemic complications. The comparison drug Mexidol increases the survival rate of animals by only 21.5%.

Example 5

Neuroprotective properties of dibenzofuranone oxime derivatives in a model of intracerebral post-traumatic hematoma, a model of hemorrhagic stroke.

The experiments were carried out on b / and male rats weighing 240-260 g. Before the operations, the rats were anesthetized with chloral hydrates (350 mg / kg). Each group contained 9 to 14 animals. Modeling of intracerebral post-traumatic hematoma - hemorrhagic stroke (HI) in rats was carried out according to the "Guidelines for conducting preclinical studies" (Publication of the Federal State Budgetary Institution "NTsEMSP" of the Ministry of Health and Social Development of Russia, Moscow, 2012, Part 1: Chapter 28; Mirzoyan R.S. et al., 2012; Voronina T.A. et al., 2012b). Using a drill in stereotaxis, craniotomy was performed in the area of the capsula interna according to the coordinates of the Buresh atlas for outbred rats - L = 3.5 mm, A = 2 mm from bregma. Using a special device (mandrin-knife), the destruction of brain tissue was carried out at a depth of H = 4.5-5.0 mm, followed by the introduction of blood taken from under the tongue of the operated animal in a volume of 0.02-0.03 ml into the site of injury. ... Sham-operated animals (LO) underwent scalping and craniotomy without destruction of brain tissue. Compounds 1a, 1b, 1c, 1e at a dose of 10 mg / kg were administered to groups of animals 2-3 hours after stroke and then once a day for the next 6 days. The substances were dissolved in physiological solution with Tween-80. The control groups with GI (No. 1, 2, 3) and LO were injected with saline with Tween-80 in an equivalent volume. Survival of animals after surgery (GI) and the development of neurological deficits (Stroke-index McGrow modified by Ganushkina (Ganushkina, 1998)) were recorded for 21 days: 1- (after 24 hours), 3-, 7-, 14- and 21- e day after surgery.

On the 1st day after the operation, in the control groups of rats with HI (No. 1,2,3), among severe neurological disorders, there were paresis of the limbs (up to 16.7%) and circling movements (up to 50%) of rats, assessed visually and by inability animals are held on a rotating rod at a speed of 10 rpm. Among mild symptoms, there were one- and two-sided half-toses (8.3-18.2%), lethargy and slowness of movements (63.6-83%) and limb weakness (45.5-66.7%), assessed by the inability of animals pull up on the crossbar (table 8). The increase in neurological deficit on the 14th day of registration was characterized by an increase in the proportion of animals with paresis, paralysis of the limbs (up to 33%) and arena movements (up to 66%) (Table 9). With the development of neurological deficit in the control groups of animals with GI, an increase in death was noted on critical days. post-stroke dynamics (on the 7th, 14th and 21st days). By the end of the experiment, 25% of rats survived in the control group with GI No. 1, and 50% in groups No. 2 and 3 (Table 7).

It was found that compound 1а at a dose of 10 mg / kg with a single administration 3 hours after hemorrhagic stroke (HI) had a positive effect on the manifestations of neurological deficit (according to the Stroke-index McGrow scale), reducing the number of animals with circus movements (by 50%) and limb weakness (by 36.7%) on the 1st day of registration after surgery relative to the corresponding values of the control group No. 1 with GI (Table 8). Compound 1a, when re-administered for 6 days, protected the animals from further death, which was recorded by a decrease in the number of dead rats on the critical days of post-stroke dynamics - 7th (by 20%) and 14th days (by 35%) (table 7). Compound 1b weakened the development of post-hemorrhagic neurological complications within 14 days after surgery: by the 14th day, no neurological disorders were observed in the surviving animals. Compound 1b reduced the death of animals on the 7th, 14th and 21st days of registration, which contributed to an increase in the survival rate by 18%, 35% and 29% of rats relative to the corresponding values of the control group with GI No. 2 (table 7). Compounds 1e and 1c did not contribute to the survival of animals after modeling hemorrhagic stroke and statistically significant weakening of neurological disorders relative to the corresponding values of control groups No. 2 and 3 (table 7, 8, 9).

Notes: GI-hemorrhagic stroke; LO-falsely operated. # - a tendency towards the reliability of values in comparison with the control group with GI, at p≤0.1 (Fisher's exact test).

Notes: GI - hemorrhagic stroke; LO-falsely operated; p / ptosis - half-ptosis. * - reliability of values in comparison with the control group, at p≤0.05 (Fisher's exact test); # - a tendency towards the reliability of values in comparison with the control group with GI, at p≤0.05 (Fisher's exact test).

Notes: GI-hemorrhagic stroke; LO-falsely operated; p / ptosis - half-ptosis. * - reliability of values in comparison with the control group with GI, at p≤0.05 (Fisher's exact test).

Thus, compounds 1a (10 mg / kg for 7 days of administration) and 1b (10 mg / kg for 7 days of administration) have a pronounced neuroprotective effect in the hemorrhagic stroke model, weaken neurological deficits, and contribute to an increase (by 35%) in the survival rate of rats by 14 m days of registration. Compounds 1c and 1e, when administered for 7 days at a dose of 10 mg / kg, did not protect against death of rats after hemorrhagic stroke.

The studies performed made it possible to establish that new compounds of dibenzofuranone oxime derivatives have anticonvulsant activity and are able to prevent generalized tonic-clonic seizures caused by MES and group clonic seizures and death caused by corazole. In the MES antagonism test, compounds 1a, 1b, 1c, 1d, 1e, 1f, and 1h, with a single administration (i / b) in the dose range from 20 to 100 mg / kg, prevent the tonic phase of convulsive manifestations and protect against death, increasing the survival rate of animals. maximum up to 66.5%. In the test of antagonism with corazole, the protective effect against death after administration of the convulsant was observed for compound 1e. Anticonvulsant activity in both tests is observed for compounds 1e. Compound 1a demonstrates antihypoxic activity under conditions of normobaric hypoxia with hypercapnia test. Compounds 1a (10 mg / kg for 7 days of administration) and 1b (10 mg / kg / 7 days of administration) have a pronounced neuroprotective effect in models of ischemic and hemorrhagic stroke, in the groups of rats of which the survival rate of animals increased and the neurological deficit was weakened. The reference drug Mexidol in the model of ischemic stroke increases the survival rate of animals by 21.5%.

Claims (12)

1. Derivatives of O-oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -one of the general formula

, Where

2. Compounds according to claim 1, having anticonvulsant and neuroprotective effects, as well as combining anticonvulsant and neuroprotective activities. 3. The use of derivatives of O-oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -one of the general formula

, Where

as agents with anticonvulsant and neuroprotective effects, as well as combining anticonvulsant and neuroprotective activities. 4. The method of obtaining compounds according to PP. 1 and 3, which consists in the interaction of the oxime 3,4,6,7,8,9-hexahydrodibenzo [b, d] furan-1 (2H) -one with acid chlorides of the corresponding acids in the presence of triethylamine. 5. A method of treating epilepsy and paroxysmal conditions, as well as post-stroke complications by introducing effective amounts of compounds according to PP. 1 and 3.