RU2748533C2 - New pheospheride derivatives with antitumor activity, method for obtaining and application of these compounds - Google Patents

Abstract

FIELD: biotechnology.SUBSTANCE: invention relates to pheospheride derivatives of general Formula I, wherein R = NR1R2, wherein R1, R2are either the same or independently selected from the group: hydrogen, AlK, wherein AlK is a methyl, ethyl, butyl, in which one hydrogen atom can be optionally replaced by a radical selected from the group: hydroxy, phenyl, vinyl, or R = О-R2, wherein R2is AlK, wherein AlK is ethyl, propyl, in which one hydrogen atom can be optionally replaced by a radical selected from the group: morpholino, piperidino, pyrrolidino, or R = S-R2, wherein R2is AlK, wherein AlK is ethyl, in which one hydrogen atom can be optionally replaced by a radical selected from hydroxy, or R is a cyclic saturated amine selected from the group: hydroxypiperidine, as well as piperidine, pyrrolidine, hydroxypiperidine, in the structure of which at least one of the hydrogen atoms is replaced by pyrrolidine. The invention also relates to a method for obtaining derivatives of Formula I.EFFECT: new compounds have been obtained that can be used in medicine as antitumor agents.6 cl, 1 tbl, 15 exI

Description

The invention relates to the field of medicine and pharmacy. Namely, to new derivatives of pheosferide - biologically active substances that have antitumor activity, as well as a method for the production and use of new derivatives of pheosferide.

Cancer is a serious and dangerous disease that is a medical and social problem all over the world. According to statistics, oncology is one of the leading causes of death in the world: the percentage of deaths from cancer is 20%.

According to WHO (World Health Organization) forecasts, over 20 years, mortality and morbidity rates from cancer will double: the number of new cases of the disease will grow from 10 million to 20 million, and the mortality rate from 6 million to 12 million. there are more than 10 million new cancer cases and more than 6 million cancer deaths.

The classic cancer treatments are cytostatic chemotherapy drugs that target nucleic acids and signaling pathways that regulate cell proliferation. The effectiveness of traditional methods of treatment (surgical, chemotherapy and radiation therapy) has not increased in recent years, which leads to the search for new approaches.

Searching for targeted cancer treatments is a priority. United by the general principle of molecular-targeted action, targeted drugs have the main targets of tissue-specific hormones and their receptors, vascular growth factors, signal transduction proteins, cell cycle regulators, and various kinases.

Thus, there is an urgent need to create new drugs for the treatment of oncological diseases, including those with high antitumor activity, but at the same time, characterized by a low degree of toxicity.

Also, preparations of herbal and natural origin have long been included in the practice of treating tumor diseases. These drugs have different chemical structures, similar mechanisms of action, but differ in the spectrum of antitumor activity and especially in side effects. Some of these drugs have a unique mechanism of action that differs from the known cytotoxic plant alkaloids. As a rule, this group of compounds exhibits relatively low toxicity.

It is known that some metabolites of natural fungi exhibit anticancer activity. For example, many micromycetes are capable of producing compounds with antitumor activity (scientific publications Tsuruo et al., 1985; Strobel, 2003; Rai et al., 2009; Kharwar et al., 2011; Evidente et al., 2013; Kornienko et al. ., 2015).

One of them is Phaeosphaeride A, a compound that has antitumor activity, selectively inhibiting the activity of one of the transcription factors, the STAT3 protein. (Phaeosphaeride A, an Inhibitor of STAT3-Dependent Signaling Isolated from an Endophytic Fungus, Katherine N. Maloney, Wenshan Hao, Jun Xu, Jay Gibbons, John Hucul, Deborah Roll, Sean F. Brady, Frank C. Schroeder, and Jon Clardy , 2006).

The structure of natural pheosferide A

Pheosferide A, a natural bicyclic substance, is considered as a platform for the creation of new anticancer drugs (1. MK Maloney, W. Hao, J. Xu, J. Gibbons, J. Hucul, D. Roll, SF Brady, FC Schroeder, J. Clardy, Org. Lett., 2006; 2. VV Abzianidze, DS Prokofieva, LA Chisty, KP Bolshakova, AO Berestetskiy, TL Panikorovskii, AS Bogachenkov and AA Holder, Bioorg. Med. Chem. Lett., 2015).

Natural pheosferide A, which has herbicidal and antitumor activity, was first isolated in 2006 from extracts of the endophytic fungus Phaeosphaeria avenaria 39 [Maloney, et al., 2006].

Pheosferide A was also isolated from the endophytic fungus cf Paraphoma sp No. 19 by the scientific group of A.O. Berestetsky (VIZR, St. Petersburg). Patent No. 2596928 STAMM Paraphoma sp. - PRODUCER FEOSPHERIDA (Federal State Budgetary Scientific Institution "All-Russian Research Institute of Plant Protection", authors: Berestetsky Alexander Olegovich, Poluektova Ekaterina Viktorovna, Sokornova Sofya Valerievna, Tokarev Yuri Sergeevich, application No. 2015140209/10, 21.09.2015).

Also, this compound was synthesized (Synthesis of the proposed structure of phaeosphaeride A, Kenichi Kobayashi, Iwao Okamoto, Nobuyoshi Morita, Tamiko Kiyotani and Osamu Tamura, 2011).

In 2015, using the method of X-ray crystallography, the molecular structure of natural phaeosphaeride A was established (Crystal structure of natural phaeosphaeride A (Victoria V. Abzianidze, Ekaterina V. Poluektova, Ksenia P. Bolshakova, Taras L. Panikorovskii, Alexander S. Bogachenkov and Alexander O. Berestetskiy, 2015), which made it possible to significantly speed up the study of the activity of this compound and open the way for its further derivatization.

Pheosferide A has a bicyclic structure with three adjacent stereocenters in its dihydropyran ring. Also known is the stereoisomer of pheosferide A - the compound pheosferides B. But, according to (1. MK Maloney, W. Hao, J. Xu, J. Gibbons, J. Hucul, D. Roll, SF Brady, FC Schroeder, J. Clardy, Org. Lett., 2006, 2. Establishment of Relative and Absolute Configurations of Phaeosphaeride A: Total Synthesis of ent-Phaeosphaeride A, Kenichi Kobayashi, Yukiko Kobayashi, Misato Nakamura, Osamu Tamura, and Hiroshi Kogen), it does not exhibit biologically active properties connections.

STAT3, a signaling protein and transcriptional activator from the STAT family of proteins, contributes to oncogenesis, therefore it attracts great interest as a target for anticancer drugs. Regulates cell proliferation, differentiation and survival. Unactivated STAT3 is usually located in the cytoplasm. After phosphorylation by Janus kinase (JAK), STAT3 dimerizes, is translated into the nucleus, then, with the participation of other protein factors, induces the transcription of those genes that should be induced by this cytokine. Atypical STAT3 activation is often found in various types of tumor cells, resulting in resistance to apoptosis and tumor cell proliferation through increased expression of genes encoding proteins such as Bcl-2, Bcl-xL, and cyclin D1.

Research shows that various STAT3 inhibitors, including synthetic small molecules and natural products, are being evaluated as promising anticancer chemotherapeutic agents.

In (Phaeosphaeride A, an Inhibitor of STAT3-Dependent Signaling Isolated from an Endophytic Fungus, Katherine N. Maloney, Wenshan Hao, Jun Xu, Jay Gibbons, John Hucul, Deborah Roll, Sean F. Brady, Frank C. Schroeder, and Jon Clardy, 2006), it was shown that pheosferide A has antitumor activity, selectively inhibiting the activity of the STAT3 protein at a concentration of IC ₅₀ = 6.1 × 10 ^-4 M.

The promising biological activity of phaeosphaeride A, as well as its unique molecular structure, has attracted a lot of attention from the synthetic community. In addition, studies of the structure-activity (SAR) inhibitory activity of STAT3 are important for potential anti-cancer therapy.

Further research and study of the substance - natural phaeosphaeride A showed that the most promising can be derivatization of this compound to obtain new substances with more pronounced antitumor, anticancer activity, and lower toxicity (Synthesis of natural phaeosphaeride A and semi-natural phaeosphaeride B derivatives, Victoria V. Abzianidze; Ksenia P. Efimova; Ekaterina V. Poluektova; Yuri G. Trishin; Victor A. Kuznetsov, 2017).

Thus, there is a need for studies of fungal metabolites and the synthesis of new substances based on them, as well as the development of new pharmaceutical preparations of these substances with antitumor properties.

The task is solved by obtaining new compounds - pheosferide derivatives with pronounced antitumor properties.

New derivatives of pheosferide, general formula I:

where R = R_one-N – R₂, where R_one,R₂either the same or independently selected from the group: hydrogen, AlK, where AlK is a group (CH₂)_nwhere n = 1-6_,in which one or more hydrogen atoms not necessary may be substituted by a radical selected from the group:

_C1-6 alkyl, hydroxy, hydroxyalkyl, benzyl, vinyl, allyl, morpholino, piperidino, pyrrolidino, hydroxypiperidino, dimethylamino, diethylamino,

or R = O-R_2,where R₂is AlK, where AlK is a group (CH₂)_nwhere n = 1-6_,in which one or more hydrogen atoms not necessary may be substituted by a radical selected from the group: C_1-6 alkyl, hydroxy, hydroxyalkyl, benzyl, vinyl, allyl, morpholino, piperidino, pyrrolidino, hydroxypiperidino, dimethylamino, diethylamino

or R = S-R_2,where R₂is AlK, where AlK is a group (CH₂)_nwhere n = 1-6_, in which one or more hydrogen atoms not necessary may be substituted by a radical selected from the group: C_1-6 alkyl, hydroxy, hydroxyalkyl, benzyl, vinyl, allyl, morpholino, piperidino, pyrrolidino, hydroxypiperidino, dimethylamino, diethylamino

or R - is a cyclic saturated amine selected from the group: morpholine, piperidine, pyrrolidine, hydroxypiperidine, as well as any of these amines, in the structure of which at least one of the hydrogen atoms is replaced by an alkyl group, hydroxyalkane, pyrrolidine, morpholine, piperidine, hydroxypiperidine.

The invention also relates to a process for the preparation of pheosferide A derivatives

Earlier, attempts were made to obtain derivatives of pheosferide A compounds by interaction of pheosperide A with a chloroacetoxy derivative (Synthesis of natural phaeosphaeride A and semi-natural phaeosphaeride B derivatives, Victoria V. Abzianidze; Ksenia P. Efimova; Ekaterina V. Poluektova; Yuri G. Trishin; Victor A. Kuznetsov, 2017). The disadvantage of this method is the very low yield of compounds.

Derivatives of pheosferide A according to the invention are obtained by reacting pheosferide A or pheosferide B in the presence of sulfochlorides with compounds selected from the group: primary amines, secondary amines, cyclic amines, alcohols, thiols.

Said sulfochlorides in the process according to the invention are compounds of the general formula R ₃ SO ₂ Cl, where R ₃ is methyl, benzyl, tolyl, triflyl groups. In particular, the following compounds can be represented: methanesulfochloride, trifluoromethanesulfochloride, benzenesulfochloride, toluenesulfochloride.

Primary amines, secondary amines, cyclic amines, alcohols, thiols in the method according to the invention are compounds of the general formula R_one-N-R₂, R_one-O-R_2,R_one-S-R_2,where where R_one,R₂either the same or independently selected from the group: hydrogen, AlK, where AlK is a group (CH₂)_nwhere n = 1-6_, in which one or more hydrogen atoms not necessary may be substituted by a radical selected from the group: C_1-6 alkyl, hydroxy, hydroxyalkyl, benzyl, vinyl, allyl, morpholino, piperidino, pyrrolidino, hydroxypiperidino, dimethylamino, diethylamino, and the cyclic amine is a cyclic saturated amine selected from the group: morpholine, piperidine, pyrrolidine, as well as any of these amines, in the structure of which one of the hydrogen atoms can be replaced by an alkyl group, hydroxyalkane, pyrrolidine, morpholine, piperidine, hydroxypiperidine.

The process of obtaining new derivatives of pheosferide A occurs with the formation of an intermediate sulfo-derivative pheosferide A or pheosferide B, formulas II and III, respectively:

with their subsequent interaction with compounds, which are primary amines, secondary amines, cyclic amines, alcohols, thiols.

The process of obtaining new compounds according to the invention can be carried out in the presence of organic solvents and / or catalysts. Dichloromethane, tetrahydrofuran, acetonitrile, toluene, benzene, dimethylformamide or mixtures thereof can be used as organic solvents. As catalysts - triethylamine, trimethylamine, diisopropylethylamine, potassium carbonate, sodium carbonate, cesium carbonate. The process is carried out at room, low or elevated temperature from 0 ° C to 80 ° C.

After the completion of the reaction, the isolation of the final product from the reaction mixture can be carried out using traditional methods of organic synthesis, selected depending on the composition of the mixture - by distilling off the solvent, recrystallization, reprecipitation, chromatography, etc. Washing, drying, evaporation, etc. can also be used.

In the course of pharmacobiological studies, it was shown that new derivatives of pheosferide A have a pronounced cytotoxic effect, which indicates the prospects of using these compounds as antitumor agents, in particular in anticancer therapy.

The invention is illustrated by the following examples.

Example 1.

Obtaining (2S, 3S, 4S) -3-hydroxy-6-methoxy-3-methyl-7-methylene-5-oxo-2-pentyl-2,3,4,5,6,7-hexahydropyrano [2,3 -c] pyrrol-4-yl methanesulfonate (sulfo derivative of pheosferide A, A)

PPA (0.084 mmol, 1 eq) was dissolved in methylene chloride (2 ml). The mixture was cooled in an ice bath, stirred for 25 minutes. Then, triethylamine (0.294 mmol, 3.5 eq) and methanesulfonyl chloride (0.21 mmol, 2.5 eq) were added dropwise. The reaction mixture was stirred for an hour. The residue was dissolved in methylene chloride (20 ml), washed with saturated NaHCO ₃ solution (20 ml) and saturated NaCl solution (20 ml). The organic layer was dried over Mg ₂ SO ₄ and rotary evaporated at 32 ° C. The molecular ion [M + H] ⁺ with a mass of 375.95, which corresponds to the mesyl derivative of pheosferide A with the molecular formula C ₁₆ H ₂₅ NO ₇ S _{2, was} fixed by mass chromatography. Received a yellow oil, 31 mg (98%).

Example 2.

Preparation of (2S, 3R, 4R) -3-hydroxy-6-methoxy-3-methyl-4- (methylamino) -7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c ] pyrrole-5 (2H) -one (compound 1)

Methylamine hydrochloride (0.37 mmol, 2 eq) and triethylamine (0.37 mmol, 2 eq) were added to a solution of the sulfonated PPA (0.18 mmol, 1 eq) in acetonitrile (5 ml) with the addition of 3 drops of water, and the reaction mixture was stirred at room temperature 48 hours. The mixture was concentrated in vacuo. The residue was dissolved in EtOAc (20 ml), washed with saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. The residue was purified by column chromatography (elution with methanol / dichloromethane 1-60) to give a yellow oil, 9 mg (17%). Ms [M + H ⁺ ]: 311.02.

Example 3.

Preparation of (2S, 3R, 4R) -3-hydroxy-4 - [(2-hydroxyethyl) amino] -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [ 2,3-c] pyrrol-5 (2H) -one (compound 2)

2-Aminoethanol (0.19 mmol, 2 eq) was added to a solution of the sulfonated PPA (0.094 mmol, 1 eq) in dry acetonitrile (5 ml) and the reaction mixture was stirred at room temperature for 48 hours. The mixture was concentrated in vacuo. The residue was dissolved in EtOAc (20 ml), washed with saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. The residue was purified by column chromatography (elution with methanol / dichloromethane 1-40) to give a yellow oil, 8 mg (25%). Ms [M + H ⁺ ]: 341.01.

Example 4.

Preparation of (2S, 3R, 4R) -3-hydroxy-4 - [(4-hydroxybutyl) amino] -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [ 2,3-c] pyrrol-5 (2H) -one (compound 3)

4-Amino-1-butanol (0.27 mmol, 2 eq) was added to a solution of the sulfonated PPA (0.13 mmol, 1 eq) in dry acetonitrile (5 ml) and the reaction mixture was stirred at room temperature for 48 hours. The mixture was concentrated in vacuo. The residue was dissolved in EtOAc (20 ml), washed with saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. The residue was purified by column chromatography (elution with methanol / dichloromethane 1-30) to give a yellow oil, 10 mg (20%). Ms [M + H ⁺ ]: 369.05.

Example 5.

Obtaining (2S, 3R, 4R) -4- (allylamino) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c ] pyrrol-5 (2H) -one (compound 4)

Allylamine (0.34 mmol, 2 eq) was added to a solution of the sulfonated PPA (0.17 mmol, 1 eq) in dry acetonitrile (5 ml) and the reaction mixture was stirred at room temperature for 48 hours. The mixture was concentrated in vacuo. The residue was dissolved in EtOAc (20 ml), washed with saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. The residue was purified by column chromatography (elution with methanol / dichloromethane 1-60) to give a yellow oil, 8 mg (14%). Ms [M + H ⁺ ]: 337.03.

Example 6.

Obtaining (2S, 3R, 4R) -4- (benzylamino) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c ] pyrrol-5 (2H) -one (compound 5)

Benzylamine (0.36 mmol, 2 eq) was added to a solution of the sulfonated PPA (0.17 mmol, 1 eq) in dry acetonitrile (5 ml) and the reaction mixture was stirred at room temperature for 24 hours. The mixture was concentrated in vacuo. The residue was dissolved in EtOAc (20 ml), washed with saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. The residue was purified by column chromatography (elution with methanol / dichloromethane 1-60) to give a yellow oil, 11 mg (17%). Ms [M + H ⁺ ]: 387.01.

Example 7.

Preparation of (2S, 3R, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-4- (4-pyrrolidin-1-ylpiperidin-1-yl) -3,4,6 , 7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one (compound 6)

4- (1-Pyrrolidinyl) piperidine (0.39 mmol, 2eq) was added to a solution of the sulfonated PPA (0.20 mmol, 1 eq) in absolute acetonitrile (5 ml) and the reaction mixture was stirred at room temperature for 24 hours. To the mixture was added 20 ml of saturated NaHCO ₃ solution. The resulting mixture was extracted three times with EtOAc (20 ml). The organic extracts were dried over MgSO ₄ and then concentrated in vacuo. Purify by column chromatography (elution with methanol / dichloromethane 1-10) to give a yellow oil, 9 mg (11%). Ms [M + H ⁺ ]: 434.12.

Example 8.

Preparation of (2S, 3R, 4R) -3-hydroxy-4- (4-hydroxypiperidin-1-yl) -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one (compound 7)

4-Hydroxypiperidine (0.25 mmol, 1.5 eq) and triethylamine (0.51 mmol, 3 eq) were added to a solution of the sulfonated PPA (0.17 mmol, 1 eq) in absolute acetonitrile (5 ml) and the reaction mixture was stirred at room temperature for 48 hours. To the mixture was added 20 ml of saturated NaHCO ₃ solution. The resulting mixture was extracted three times with EtOAc (20 ml). The organic extracts were dried over MgSO ₄ and then concentrated in vacuo. Purify by column chromatography (elution with methanol / dichloromethane 1-50) to give a yellow oil, 10 mg (16%). Ms [M + H ⁺ ]: 381.05.

Example 9.

Obtaining (2S, 3R, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-4-pyrrolidin-1-yl-3,4,6,7-tetrahydropyrano [2,3 -c] pyrrol-5 (2H) -one (compound 8)

Pyrrolidine (0.12 mmol, 1.5 eq) and triethylamine (0.25 mmol, 3 eq) were added to a solution of the sulfonated PPA (0.084 mmol, 1 eq) in absolute acetonitrile (5 ml) and the reaction mixture was stirred at room temperature for 48 hours. To the mixture was added 20 ml of saturated NaHCO ₃ solution. The resulting mixture was extracted three times with EtOAc (20 ml). The organic extracts were dried over MgSO ₄ and then concentrated in vacuo. Purify by column chromatography (elution with methanol / dichloromethane 1-60) to give a yellow oil, 8 mg (27%). Ms [M + H ⁺ ]: 351.04.

Example 10.

Preparation of (2S, 3S, 4R) -3-hydroxy-4 - [(2-hydroxyethyl) thio] -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [ 2,3-c] pyrrol-5 (2H) -one (compound 9)

To a suspension of 2-mercaptoethanol (0.11 mmol, 1.1 eq) and K ₂ CO ₃ (0.20 mmol, 2 eq) in acetonitrile (2 ml) at 0 ° С was added dropwise a solution of the sulfonated PPA (0.10 mmol, 1 eq) in acetonitrile ( 1 ml). The reaction mixture was allowed to warm to room temperature and stirred for 72 hours. The mixture was then hydrolyzed with ice water (20 ml) and extracted with diethyl ether (2 x 30 ml). The combined extracts were dried over MgSO ₄ and then concentrated in vacuo. Purify by column chromatography (elution with methanol / dichloromethane 1-50) to give a yellow oil, 7.2 mg (20%). Ms [M + H ⁺ ]: 357.95.

Example 11.

Preparation of (2S, 3S, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-4- (2-morpholin-4-ylethoxy) -2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one (compound 10)

To a suspension of sulfo-derivative PPA (0.18 mmol, 1 eq) in anhydrous tetrahydrofuran (2 ml), 2-morpholinoethanol (0.90 mmol, 5 eq) was added, and the mixture was stirred in a closed flask at 70 ° С for 48 h. The mixture was concentrated in vacuum. The residue was dissolved in EtOAc (20 ml), washed with saturated NaHCO ₃ solution (15 ml), saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. Purify by column chromatography (elution with methanol / dichloromethane 1-10) to give a yellow oil, 10 mg (14%). Ms [M + H ⁺ ]: 411.16.

Example 12.

Preparation of (2S, 3S, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-4- (3-piperidin-1-ylpropoxy) -3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one (compound 11)

To a suspension of sulfo-derivative PPA (0.18 mmol, 1 eq) in anhydrous tetrahydrofuran (2 ml) was added 3-piperidino-1-propanol (0.90 mmol, 5 eq), and the mixture was stirred in a closed flask at 70 ° С for 48 h. The mixture was concentrated in a vacuum. The residue was dissolved in EtOAc (20 ml), washed with saturated NaHCO ₃ solution (15 ml), saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. Purify by column chromatography (elution with methanol / dichloromethane 1-10) to give a yellow oil, 12 mg (16%). Ms [M + H +]: 423.20.

Example 13.

Preparation of (2S, 3S, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-4- (3-pyrrolidin-1-ylpropoxy) -3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one (compound 12)

To a suspension of sulfo-derivative PPA (0.18 mmol, 1 eq) in anhydrous tetrahydrofuran (2 ml) was added 3-pyrrolidino-1-propanol (0.90 mmol, 5 eq), and the mixture was stirred in a closed flask at 70 ° С for 48 h. The mixture was concentrated in a vacuum. The residue was dissolved in EtOAc (20 ml), washed with saturated NaHCO ₃ solution (15 ml), saturated NaCl solution (15 ml). The organic extract was dried over MgSO ₄ and then concentrated in vacuo. Purify by column chromatography (elution with methanol / dichloromethane 1-10) to give a yellow oil, 10 mg (14%). Ms [M + H +]: 409.22.

Example 14.

Study of the cytotoxicity of compounds 1-8.

The cytotoxicity of the products was studied on the cell lines MCF-7 (breast cancer), PC-3 (adenocarcinoma of the prostate), HCT-116 (colon cancer), A549 (lung cancer), K562 (chronic myeloid leukemia), THP -1 (acute monocytic leukemia), NCI-H929 and RPMI8226 (multiple myeloma) Jurkat (T cell lymphoblastic leukemia). Human embryonic fibroblasts (HEF) were used as a control.

The cultivation was carried out at 37 ° C in an atmosphere containing 5% CO ₂ . DMEM medium was used for MCF-7, A594, and HEP cells; the rest of the cells were cultured in RPMI 1640 medium in the presence of 10% fetal bovine serum and gentamicin at a concentration of 40 mg / ml.

The cytotoxic effect was assessed using a standard MTT test, calculating the proportion of viable cells from the results of colorimetric determination of the MTT reduction product (formazan).

The concentration of the test product providing a 50% cytotoxic effect relative to the control (IC50) was calculated using the "Program for calculating the IC50 of chemotherapeutic substances" according to the results of the analysis of cell viability in the presence of the investigated substances at 10 concentrations (from 2 * 10 ^-7 to 1 * 10 ^-4 M). Each concentration was analyzed in 3-4 repetitions.

In the process of studying the cytotoxicity of the compounds of general formula I and the reference substance (etoposide), the results shown in the table were obtained.

Table 1

Compound Adhesive cell cultures, IC50 (μM) HCT-116 PC3 MCF-7 A549 FECH PPA 44 32 twenty 41 nineteen one 4.0 4.8 4.6 13 22 2 4.6 4.5 3.9 fifteen sixteen 3 4.9 5.6 5.3 eleven fifteen four 8.4 6.3 4.1 13 five five 13 25 36 38 53 6 4.3 3.2 2,3 12 22 7 2.6 4.3 3.5 9.4 6,7 eight 0.47 0.2 4.8 10 four Etoposide 22 2.7 9.6 > 100 > 100

Compound Suspension cell cultures, IC50 (μM) K562 NCI-H929 Jurkat THP-1 RPMI8226 PPA twenty 6.5 10 nineteen 9.6 one 1.9 0.56 1.9 2.4 3.2 2 17 2,3 17 1.9 2.8 3 3.8 2.1 1.9 2.2, 2.5 four 10 2,3 3.2 3.3 6.0 five fourteen 7,7 10 fifteen sixteen 6 6.4 3.2 one 4.9 10 7 3.2 1.9 1.8 1.9 1.6 eight 0.54 0.2 0.75 2.2 0.25 Etoposide 4.5 0.9 1.4 1.0 4.8

The table shows that PPA derivatives have a cytotoxic effect comparable to that of etoposide, and in some cases, a more pronounced cytotoxic effect. Thus, for substance 8, the IC50 for multiple myeloma cell lines (RPMI8226 and NCI-H929) is several times lower than for etoposide.

Example 15.

The study of acute toxicity after intraperitoneal injection of 8 was performed on white outbred mice weighing 16-20 g. Each study group included 10 animals. Stock solution 8 was prepared using DMSO, then diluted with phosphate-buffered saline to a concentration providing the required dose of the drug in a volume not exceeding 0.1 ml. The final concentration of DMSO in the solution for administration is 5%. The dose range used in the study was 8.25 to 200 mg / kg. Survival and manifestations of toxic effects were recorded within 14 days after a single intraperitoneal injection of the drug. Animals in the control group received a solvent containing 5% DMSO. According to the results of the study, it was shown that the LD50 value exceeds 200 mg / kg. For intraperitoneal administration - the obtained value corresponds to the 3rd hazard class (moderately hazardous substances). Acute toxicity 8 is comparable to that of etoposide (LD50 for etoposide (RTECS # KC0190000) when injected intraperitoneally is 64 mg / kg).

Claims (31)

1. Derivatives of pheosferide, general formula I

where R = NR ₁ R ₂ , where R ₁ , R _{2 are} either the same or independently selected from the group: hydrogen, AlK, where AlK is methyl, ethyl, butyl, in which one hydrogen atom may optionally be substituted by a radical selected from groups: hydroxy, phenyl, vinyl, or R = O-R ₂ , where R ₂ is AlK, where AlK is ethyl, propyl, in which one hydrogen atom may optionally be replaced by a radical selected from the group: morpholino, piperidino, pyrrolidino, or R = SR ₂ , where R ₂ is AlK, where AlK is ethyl, in which one hydrogen atom may optionally be substituted with a radical selected from hydroxy, or R - is a cyclic saturated amine selected from the group: hydroxypiperidine, as well as piperidine, pyrrolidine, hydroxypiperidine, in the structure of which at least one of the hydrogen atoms is replaced by pyrrolidine. 2. Compounds according to claim 1, which, in particular, are: - (2S, 3R, 4R) -3-hydroxy-6-methoxy-3-methyl-4- (methylamino) -7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c ] pyrrole-5 (2H) -one; - (2S, 3R, 4R) -3-hydroxy-4 - [(2-hydroxyethyl) amino] -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [ 2,3-c] pyrrol-5 (2H) -one; - (2S, 3R, 4R) -3-hydroxy-4 - [(4-hydroxybutyl) amino] -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [ 2,3-c] pyrrol-5 (2H) -one; - (2S, 3R, 4R) -4- (allylamino) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c ] pyrrole-5 (2H) -one; - (2S, 3R, 4R) -4- (benzylamino) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c ] pyrrole-5 (2H) -one; - (2S, 3R, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-4- (4-pyrrolidin-1-ylpiperidin-1-yl) -3,4,6 , 7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one; - (2S, 3R, 4R) -3-hydroxy-4- (4-hydroxypiperidin-1-yl) -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one; - (2S, 3S, 4R) -3-hydroxy-4 - [(2-hydroxyethyl) thio] -6-methoxy-3-methyl-7-methylene-2-pentyl-3,4,6,7-tetrahydropyrano [ 2,3-c] pyrrol-5 (2H) -one; - (2S, 3S, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-4- (2-morpholin-4-ylethoxy) -2-pentyl-3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one; - (2S, 3S, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-4- (3-piperidin-1-ylpropoxy) -3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one; - (2S, 3S, 4R) -3-hydroxy-6-methoxy-3-methyl-7-methylene-2-pentyl-4- (3-pyrrolidin-1-ylpropoxy) -3,4,6,7-tetrahydropyrano [2,3-c] pyrrol-5 (2H) -one. 3. A method of obtaining derivatives of pheosferide of General formula I

, where R = NR ₁ R ₂ , where R ₁ , R _{2 are} either the same or independently selected from the group: hydrogen, AlK, where AlK is methyl, ethyl, butyl, in which one hydrogen atom may optionally be substituted by a radical selected from groups: hydroxy, phenyl, vinyl, or R = O-R ₂ , where R ₂ is AlK, where AlK is ethyl, propyl, in which one hydrogen atom may optionally be replaced by a radical selected from the group: morpholino, piperidino, pyrrolidino, or R = SR ₂ , where R ₂ is AlK, where AlK is ethyl, in which one hydrogen atom may optionally be substituted with a radical selected from hydroxy, or R - is a cyclic saturated amine selected from the group: pyrrolidine, hydroxypiperidine, as well as piperidine, pyrrolidine, hydroxypiperidine, in the structure of which at least one of the hydrogen atoms is replaced by pyrrolidine, by the interaction of pheosperide A or pheosferide B in the presence of at least at least one sulfochloride of the general formula R ₃ SO ₂ Cl, where R ₃ is methyl, benzyl, tolyl, trifluoromethyl groups with compounds selected from the group: primary amines, secondary amines, cyclic amines, alcohols, thiols, which are, respectively, compounds of the general formula R ₁ -NR ₂ , R ₁ -O-R ₂ , R ₁ -SR ₂ , where R ₁ , R _{2 are} either the same or independently selected from the group: hydrogen, AlK, where AlK is methyl, ethyl, butyl, in which one hydrogen atom may optionally be substituted by a radical selected from the group: hydroxy, phenyl, vinyl, or AlK represents ethyl, propyl, in which one hydrogen atom may optionally be substituted by a radical selected from and 3 groups: morpholino, piperidino, pyrrolidino, or AlK is ethyl, in which one hydrogen atom may optionally be replaced by a radical selected from hydroxy, and the cyclic amine is a cyclic saturated amine selected from the group: pyrrolidine, hydroxypiperidine, and piperidine , pyrrolidine, hydroxypiperidine, in the structure of which at least one of the hydrogen atoms is replaced by pyrrolidine, in the presence of at least one catalyst selected from the group: triethylamine, trimethylamine, diisopropylethylamine, potassium carbonate, sodium carbonate, cesium carbonate, in the presence of at least one organic solvent selected from the group: dichloromethane, tetrahydrofuran, acetonitrile, toluene, benzene, dimethylformamide, and a temperature from 0 ° C to 80 ° C. 4. The method according to p. 3, characterized in that the interaction of pheosferide A or pheospheride B in the presence of sulfochloride is the formation of an intermediate sulfo-derivative pheosferide A or pheosferide B, followed by its interaction with compounds selected from the group: primary amines, secondary amines, cyclic amines, alcohols, thiols, which are, respectively, compounds as indicated in clause 3. 5. The method according to PP. 3, 4, where the following compounds are selected as sulfochlorides: methanesulfochloride, trifluoromethanesulfochloride, benzenesulfochloride, toluenesulfochloride. 6. Application of pheosferide derivatives of general formula I

where R = NR ₁ R ₂ , where R ₁ , R _{2 are} either the same or independently selected from the group: hydrogen, AlK, where AlK is methyl, ethyl, butyl, in which one hydrogen atom may optionally be substituted by a radical selected from groups: hydroxy, phenyl, vinyl, or R = O-R ₂ , where R ₂ is AlK, where AlK is ethyl, propyl, in which one hydrogen atom may optionally be replaced by a radical selected from the group: morpholino, piperidino, pyrrolidino, or R = SR ₂ , where R ₂ is AlK, where AlK is ethyl, in which one hydrogen atom may optionally be substituted with a radical selected from hydroxy, or R - is a cyclic saturated amine selected from the group: pyrrolidine, hydroxypiperidine, as well as piperidine, pyrrolidine, hydroxypiperidine, in the structure of which at least one of the hydrogen atoms is replaced by pyrrolidine, as antitumor agents.