RU2695062C2 - Novel 3,5-divinyl-pyrazole derivatives for medical use - Google Patents

Abstract

FIELD: medicine; pharmacology.

SUBSTANCE: invention refers to pharmacology, and concerns new chemical compounds of general formula (I) or their pharmaceutically acceptable salts which are inhibitors of P-glycoprotein. In formula (I) R₁, R₂, R₃, R₄ independently represent H, halogen, NO₂; R_x - is selected from -C(=S)NHR^A, -C(=O)OR^A, -S(=O)₂NR^AR^B, -C(=O) NR^AR^B, 5-membered partially or completely unsaturated heterocycle containing 2 N atoms; R^A is independently selected and represents C_1-6-alkyl, partially or completely halogenated C_1-8-alkyl; R^B is independently selected and represents H, C_1-8-alkyl or partially or completely halogenated C_1-8-alkyl; alternatively, in case R_x is -C (= O) NR^AR^B, R^A and R^B together with nitrogen atom, to which they are attached, can form saturated cycle containing 5 atoms; R_p is independently selected and represents C_1-6-alkyl, partially or completely halogenated C_1-6-alkyl; R_m is selected independently and is H, C_1-6-alkyl, partially or completely halogenated C_1-8-alkyl; alternatively neighbouring groups R_p and R_m together with two oxygen atoms, to which they are attached, and two carbon atoms of the benzene ring can form a non-aromatic ring containing 5 to 6 atoms. Given chemical compounds can be used to suppress multiple drug resistance in the patients receiving the various drugs for a long time. Invention also relates to the use of the above compounds and a pharmaceutical composition containing said compounds.

(I).

EFFECT: these compounds can be used in therapy of central nervous system diseases, infectious, allergic, autoimmune, oncological and other diseases.

9 cl

Description

Technical field

The invention relates to medicine, namely to pharmacology, and relates to new chemical compounds with high efficiency in inhibiting P-glycoprotein, which can be used in the treatment of diseases of the central nervous system, infectious, allergic, autoimmune, oncological and other diseases. In particular, these chemical compounds can be used to suppress multidrug resistance in patients who receive various drugs for a long time.

The level of technology

P-glycoprotein (P-gp) belongs to the family of ABC carriers (ATP-binding cassette (ABC) transporters) - proteins that carry ATP-dependent transport of substances through the cell membrane against a concentration gradient [Higgins, CF, Nature, 2007. 446 ( 7137): p. 749-57]. Due to its broad substrate specificity, human P-glycoprotein is involved in the export of medicinal substances from cells, which leads to the emergence of multidrug resistance [Borst, P. and R.O. Elferink, Annu Rev Biochem, 2002. 71: p. 537-92]. Multiple drug resistance is the immunity of cells simultaneously to a variety of chemotherapeutic drugs of different chemical structure. In addition, since specific ABC transporters are expressed in pathogenic microorganisms, providing resistance to antibiotics, inhibition of bacterial ABC transporters can increase the effectiveness of antimicrobial chemotherapy [Ouellette, M., D. Legare, and B. Papadopoulou, J Mol Microbiol Biotechnol, 2001 3 (2): p. 201-6]. P-gp is actively expressed in the endothelium of brain capillaries and plays a key role in regulating the permeability of the blood-brain barrier for drugs and xenotoxins [Schinkel, A.H., Adv Drug Deliv Rev, 1999. 36 (2-3): p. 179-194].

The active study of P-glycoprotein has been going on for more than 30 years and, apparently, P-glycoprotein is the most well-studied protein from the family of ABC carriers. During this time, a greater number of various synthetic and natural compounds have been found that can inhibit P-glycoprotein, leading to an increase in the penetration of drugs into the cell and the suppression of multidrug resistance [Shukla, S., C.P. Wu, and S.V. Ambudkar, Expert Opin Drug Metab Toxicol, 2008. 4 (2): p. 205-23].

Three generations of P-glycoprotein inhibitors are currently known: most of the first generation P-gp inhibitors were developed before 1990 and had their own pharmacological activity (for example, Verapamil [US2002016565120 (2002)], Cyclosporin [US200710060635A1 (2007), US20090088393 (2009) , US200510288222A1 (2005)], Quinidine, Reserpine, etc.), however, the possibility of their use for the suppression of multidrug resistance was limited by the high toxicity of these compounds. To eliminate the problems associated with the toxicity of compounds of the first generation, P-gp inhibitors of the second generation were proposed (for example, D-isomer Verapamil, which had significantly less cardiotoxicity compared to the racemate) [Krishna, R. and L.D. Mayer, Eur J Pharm Sci, 2000. 11 (4): p. 265-83]. At the same time, second-generation P-gp inhibitors had low selectivity for other proteins from the ABC transporter family and due to this in some cases led to the development of serious side effects [Thomas, H. and H.M. Coley, Cancer Control, 2003. 10 (2): p. 159-65; Boesch, D., et al., Cancer Res, 1991. 51 (16): p. 4226-33]. An intensive search for new, more specific and low-toxic P-gp inhibitors led to the development of a third generation of compounds. Third Generation P-gp Inhibitors (Elakridar [Van Tellingen, OZ, NL, Methods and Kanamura, K., et al., Nucl Med Biol, 2009. 36 (3): p. 239-46], Tarikvidar [Pajeva, IK and M. Wiese, AAPS J, 2009. 11 (3): p. 435-44], Zosikvidar [Sato, W., et al., Cancer Res, 1991. 51 (9): p. 2420-4]) significantly superior to inhibitors of the first and second generation, both in their affinity for P-glycoprotein and in selectivity for other ABC transporters. At the same time, clinical trials of most third-generation P-gp inhibitors have been discontinued, either due to lack of efficacy, or due to serious side effects.

Many natural compounds, such as curcumin and flavonoids (for example, Kempferol and Quercetin), are able to inhibit P-glycoprotein and suppress multidrug resistance [Limtrakul, P., O. Khantamat, and K. Pintha, J Chemother, 2005. 17 (1 ): p. 86-95]. The closest analogue of the compounds of this invention is Curcumin I. Curcumin (a mixture of curcuminoids isolated from turmeric powder), widely used as a spice, dye and therapeutic agent in oriental medicine, has inhibitory activity against P-glycoprotein [Chearwae, W., et al. , Biochem Pharmacol, 2004. 68 (10): p. 2043-52]. Curcumin I (present in the mixture at a concentration of> 70%) is the most active inhibitor of P-gp. At the same time, Curcumin I has several disadvantages - relatively low affinity for P-gp (according to various data from 1 to 10 uM), low bioavailability when administered orally and rapid metabolism.

Thus, the creation of new compounds capable of inhibiting P-glycoprotein, with good pharmacokinetic parameters and not causing serious side effects, is a practically important task.

This invention relates to a new group of chemical compounds with increased efficacy in inhibiting P-glycoprotein, as well as high affinity for P-glycoprotein and high bio-insensitivity, and promising for the suppression of multi-drug resistance caused by the activity of P-glycoprotein.

DISCLOSURE OF INVENTION

The task (technical result) of the present invention is the development of new chemical compounds with high efficiency in inhibiting P-glycoprotein, as well as having high bioavailability and affinity for P-glycoprotein, and promising for use in the treatment of diseases in the development mechanism of which P-glycoprotein is involved in particular, by suppressing multidrug resistance or to increase the absorption or distribution of the drug. Such diseases can be diseases of the central nervous system, infectious, allergic, autoimmune, oncological diseases.

This technical result is achieved by obtaining compounds of the general formula (I) or their pharmaceutically acceptable salts, hydrates or solvates:

Where:

R ₁ , R ₂ , R ₃ , R ₄ - are chosen independently and represent H, halogen, OH,

C _1-6 -alkoxy, NO ₂ , -C (= S) NHR ^A , -C (= O) OR ^A , -S (= O) ₂ OR ^A , -C (= S) OR ^A , -S ( = O) ₂ NHR ^A , -C (= O) NHR ^A , -S (= O) ₂ NR ^A R ^B or -C (= O) NR ^A R ^B ;

R _X is selected from the group —C (= S) NHR ^A , —C (= O) OR ^A , —S (= O) ₂ OR ^B , —C (= S) OR ^B ,

-S (= O) ₂ NHR ^A , -C (= O) NHR ^A , -S (= O) ₂ NR ^A R ^B , -C (= O) NR ^A R ^B , 5-6-membered partially or completely unsaturated heterocycle containing 1 to 2 N atoms;

R ^A is independently selected and represents C _1-6 alkyl, C _1-8 alkoxy, partially or fully halogenated C _1-8 alkyl or C _5-6 cycloalkyl;

R ^B is independently selected and is H, C _1-8 alkyl, C _1-8 alkoxy, or partially or fully halogenated C _1-8 alkyl;

alternatively, in cases where any of R ₁ , R ₂ , R ₃ , R ₄ , R _X is

-S (= O) ₂ NR ^A R ^B or- ^C (= O) NR ^A R ^B , R ^A and R ^B , together with the nitrogen atom to which they are attached, can form a saturated cycle containing from 5 to 6 atoms ;

R _p - is independently selected and represents a C ₁ - ₆ -alkyl, partially or completely halogenated C _1-6 alkyl;

R _m - is independently selected and represents H, C ₁ - ₆ -alkyl, partially or completely halogenated C _1-8 alkyl;

alternatively, the adjacent R _p and R _m groups together with the two oxygen atoms to which they are attached, and the two carbon atoms of the benzene ring, can form a non-aromatic cycle containing from 5 to 6 atoms.

The chemical compounds that are the subject of the present invention are characterized by a concentration of half-maximal inhibition of P-glycoprotein in the range from 0.1 to 5000 nM.

Preferred embodiments of the invention

A separate class of compounds of interest includes compounds of formula (I) in which:

R ₁ , R ₂ , R ₃ , R ₄ - are chosen independently and represent H, halogen, NO ₂ ;

R _X - represents -C (= S) NHR ^A , -C (= O) OR ^A , -C (= S) OR ^B , -C (= O) NHR ^A ,

-S (= O) ₂ NR ^A R ^B , -C (= O) NR ^A R ^B or a 5-6-membered partially or fully unsaturated heterocycle containing 1 to 2 N atoms;

R ^A - is chosen independently and represents a C _1-6 -alkyl, C _1-8 -alkoxy, partially or fully halogenated C _1-8 -alkyl;

R ^B - is chosen independently and represents H, C _1-8 -alkyl, C _1-8 -alkoxy, partially or fully halogenated C _1-8 -alkyl;

alternatively, in cases where R _X is —C (═O) NR ^A R ^B , R ^A and R ^B together with the nitrogen atom to which they are attached, may form a saturated cycle containing from 5 to 6 atoms;

R _p is independently selected and represents CH ₃ , partially or fully halogenated C _1-6 alkyl;

R _m is selected independently and represents H, CH ₃ , partially or fully halogenated C _1-8 -alkyl.

Another separate class of compounds of interest includes compounds of formula (I) in which:

R ₁ , R ₂ , R ₃ , R ₄ - are chosen independently and represent H, halogen, NO ₂ ;

R _X - represents -C (= S) NHR ^A , -C (= O) OR ^A , -C (= S) OR ^B , -C (= O) NHR ^A ,

-S (= O) ₂ NR ^A R ^B , -C (= O) NR ^A R ^B , 5-6-membered partially or fully unsaturated heterocycle containing 1 to 2 N atoms;

R ^A - is chosen independently and represents a C _1-6 -alkyl, C _1-8 -alkoxy, partially or fully halogenated C _1-8 -alkyl;

R ^B - is chosen independently and represents H, C _1-8 -alkyl, C _1-8 -alkoxy, partially or fully halogenated C _1-8 -alkyl;

alternatively, in cases where R _X is —C (═O) NR ^A R ^B , R ^A and R ^B together with the nitrogen atom to which they are attached, may form a saturated cycle containing from 5 to 6 atoms;

R _p - is independently selected and represents a CH _3, partially or completely halogenated C ₁ - ₆ -alkyl;

R _m is selected independently and represents H, CH ₃ , partially or fully halogenated C ₁ - ₈ -alkyl,

however, the neighboring groups R _p and R _m together with the two oxygen atoms to which they are attached, and the two carbon atoms of the benzene ring, form a non-aromatic cycle containing from 5 to 6 atoms.

Illustrative examples of a separate subclass of compounds of interest include compounds of formula (I) in which R m is CH ₃ :

Illustrative examples of a separate subclass of compounds of interest include compounds of formula (I) in which Rp and Rm together with the two oxygen atoms to which they are attached and two carbon atoms of the benzene ring form a non-aromatic cycle:

Illustrative examples of a separate subclass of compounds of interest include compounds of formula (I) in which Rp and / or Rm are partially or fully halogenated C _1-6 alkyl:

Illustrative examples of a particular subclass of compounds of interest include compounds of formula (I) in which R ₁ and / or R ₂ and / or R ₃ and / or R ₄ are H or halogen:

Illustrative examples of a separate subclass of compounds of interest include compounds of formula (I) in which R _X is —C (═O) OR ^A ,

—C (═O) NHR ^A , —C (═O) NR ^A R ^B , i.e., substituted carboxylic ester or amide:

Illustrative examples of a separate subclass of compounds of interest include compounds of formula (I) in which R _X is

-C (= S) NHR ^A , -C (= S) OR ^B , i.e., substituted thio-carboxylic acid ester or amide. Illustrative examples of this class are the following compounds:

Illustrative examples of a separate subclass of compounds of interest include compounds of formula (I) in which R _X is a substituted imidazole, imidazolyl, or sulfonamide. Illustrative examples of this class are the following compounds:

Specific examples of the compounds presented in the description should be considered as illustrative and not as restrictive. It should be clear that it is possible to implement various modifications without departing from the gist of the present invention.

This invention also relates to the use of the compounds of the invention as inhibitors of P-glycoprotein.

The present invention also relates to the use of the compounds of the invention for the preparation of a pharmaceutical composition for the treatment of diseases associated with the activity of P-glycoprotein.

In particular, such pharmaceutical compositions can be used to suppress multidrug resistance caused by P-glycoprotein activity.

In addition, the invention provides for the use of compounds according to the invention for the preparation of a pharmaceutical composition that can be used to increase the bioavailability of various oral drugs with low intestinal adsorption, by suppressing the activity of P-glycoprotein in the cells of the digestive tract of humans or animals.

The invention also involves the use of compounds that are the subject of the invention, to obtain a pharmaceutical composition for increasing the concentration of drugs, for example, antiepileptic drugs, in the brain cells of humans or animals.

The invention also involves the use of compounds that are the subject of the invention, to obtain a pharmaceutical composition to increase the concentration of cytostatic and / or cytotoxic drugs in tumor cells. In particular, such cytostatic or cytotoxic drugs can be doxorubicin, daunorubicin, epirubicin, idarubicin, teniposide, methotrexate, pemetrexed, etoposide.

The invention also encompasses pharmaceutical compositions containing the compounds of the present invention, including compounds of any of the classes or subclasses described, including any of the formulas described above, in a therapeutically effective amount, in combination with at least one therapeutically acceptable carrier, is solvent and / or filler.

In addition, the invention provides the pharmaceutical compositions described above, which further include a chemotherapeutic or pharmacotherapeutic agent.

In particular, the chemotherapeutic agent may be etoposide, doxorubicin, vinblastine, ticarcillin, carbpenicillin, mezlocillin, azlocillin, ceftbiprol, ceftaroline, ceftlozan, methotrexate, and others.

The pharmacotherapeutic agent, in particular, can be digoxin, dexamethasone and others.

The invention also includes the preparation of compounds of formula (I) or any other compounds of the present invention.

Definitions

The following definitions apply herein unless expressly stated otherwise. In addition, unless otherwise indicated, all occurrences of functional groups are selected independently.

The term "alkyl" by itself, or as part of another substituent, refers to straight or branched saturated hydrocarbon groups, including hydrocarbon groups having a specified number of carbon atoms (i.e., C _1-6 means from one to six carbon atoms). For example, "alkyl" can mean methyl, ethyl, n-propyl, isopropyl, cyclopropyl, butyl, isobutyl, sec-butyl, cyclobutyl, tert-butyl, cyclobutyl, pentyl, cyclopentyl, tert-pentyl, isopentyl, hexyl, isohexyl, cyclohexyl etc. As an illustration, substituted alkyl groups include, but are not limited to, the following groups: fluoromethyl, difluoromethyl, trifluoromethyl, 2-fluoroethyl, 3-fluoropropyl, hydroxymethyl, 2-hydroxyethyl, 3-hydroxypropyl, etc.

The term "alkoxy" refers to alkyl groups, as defined above, and which are attached to the molecule via a bridging oxygen atom. For example, the term "C _1-8 alkoxy" means -O-alkyl, where the alkyl group contains from 1 to 8 carbon atoms in the form of a linear (unbranched) or branched chain or in the form of a cycle. By way of illustration, alkoxy groups include, but are not limited to, the following groups: methoxy, ethoxy, n-propoxy, n-butoxy, tert-butoxy, allyloxy, cyclobutoxy, etc.

The term "halogen" by itself or in part of another term refers to a fluorine, chlorine, bromine or iodine atom.

The term "halogenated alkyl" includes branched and linear saturated hydrocarbon chains in which one or more hydrogen atoms are replaced by halogen. Examples of halogenated alkyl groups include, but are not limited to, the following groups: difluoromethyl, trifluoromethyl, trichloromethyl, pentafluoroethyl, 1,1,1,3,3,3-hexafluoro-2-methylpropan-2-yl, and the like.

The term “cycloalkyl” means an alicyclic saturated group having the indicated number of carbon atoms.

The term "non-aromatic cycle" refers to saturated or partially unsaturated cycles.

The term "heterocycle" ("heterocyclic"), in the context of the invention, means a cyclic system having from 5 to 6 atoms, which, along with carbon, also include atoms of other elements. Such heteroatoms of the invention may be N or O. The term “heterocycle” refers to saturated, partially unsaturated, or aromatic rings (heteroaryl). The degree of saturation of the heterocycle, as well as the number of heteroatoms are indicated separately.

This invention contains only such combinations of substituents and derivatives, which form a stable or chemically possible compound. A stable or chemically feasible compound is a compound whose stability is sufficient for its synthesis and analytical detection. Preferred compounds of this invention are fairly stable and do not decompose at temperatures up to 40 ° C in the absence of chemically active conditions for at least one week.

The implementation of the invention

The compounds that are the subject of the present invention, can be obtained using various well-known synthetic methods, including using the methods described below. The synthesis of chemical compounds that are the subject of the present invention can be carried out from commercially available initial reagents or initial reagents, which can be obtained by the methods described in works known from the prior art (Weigand - Hilgetag. Methods of experiment in organic chemistry. Ed. Prof. N. N. Suvorov. M., Chemistry, 1968, "Synthesis of heterocyclic compounds" Vol. 1-16 Yerevan 1956-1987; "Synthesis of organic preparations" Part 1-12 M. 1949-1964; "Synthesis of organic compounds with hydrogen isotopes "Murray A., Williams D . L. Moscow, Publisher IL, 1961;). The listed methods are not exhaustive and allow the introduction of reasonable modifications. These reactions should be carried out using suitable solvents and materials. When implementing these general methods for the synthesis of specific substances, it is necessary to take into account the functional groups present in the substances and their influence on the course of the reaction. To obtain certain substances, it is necessary to change the order of the stages or give preference to one of several alternative synthesis schemes.

General scheme of the final stage of the preparation of the compounds according to the invention

All compounds that are the subject of this invention, can be obtained on the basis set forth in the description of synthetic approaches, examples of experimental techniques and well-known methods and materials.

Examples of the synthesis of compounds according to the invention

Synthesis of intermediate compounds

Curcuminoid Synthesis I (CUR I)

To a solution of 350 mg (5.0 mmol) of boric acid anhydride in 60 ml of anhydrous ethyl acetate, 1.0 ml (10 mmol) of acetylacetone is added and the reaction mixture is stirred for 30 minutes at 40 ° C, then 3.04 g (20 mmol) of vanillin is added and 10.8 ml (40 mmol) of tributylborate. The reaction mixture is stirred for 30 minutes at 50 ° C, after which 0.4 ml (5 mmol) of n-butylamine in 5 ml of anhydrous ethyl acetate are added dropwise over 15 minutes. The reaction mixture is heated to 70 ° C and stirred at this temperature for 10 hours, 30 ml of 1N aqueous hydrochloric acid are added and stirred at 60 ° C for another 2 hours, cooled, the organic layer is separated, and the aqueous layer is extracted with ethyl acetate (3 x 50 ml). The combined organic phases are dried over anhydrous sodium sulfate and concentrated in vacuo. The residue is recrystallized from methanol and dried. Receive: 957 mg (52%) of the product.

Synthesis of Curcuminoid II (CUR II)

To a solution of 350 mg (5.0 mmol) of boric acid anhydride in 60 ml of anhydrous ethyl acetate, 1.0 ml (10 mmol) of acetylacetone is added and the reaction mixture is stirred for 30 minutes at 40 ° C, after which 3.32 g (20 mmol) of 4-ethoxy-3 is added -hydroxybenzaldehyde and 10.8 ml (40 mmol) of tributylborate. The reaction mixture is stirred for 30 minutes at 50 ° C, after which 0.4 ml (5 mmol) of n-butylamine in 5 ml of anhydrous ethyl acetate are added dropwise over 15 minutes. The reaction mixture is heated to 70 ° C and stirred at this temperature for 10 hours, 30 ml of 1N aqueous hydrochloric acid are added and stirred at 60 ° C for another 2 hours, cooled, the organic layer is separated, and the aqueous layer is extracted with ethyl acetate (3 × 50 ml). The combined organic phases are dried over anhydrous sodium sulfate and concentrated in vacuo. The residue is recrystallized from methanol and dried. Receive: 524 mg (34%) of the product.

Synthesis of Dimethylcurcumin (DMCUR)

To a solution of 350 mg (5.0 mmol) of boric acid anhydride in 60 ml of anhydrous ethyl acetate, 1.0 ml (10 mmol) of acetylacetone is added and the reaction mixture is stirred for 30 minutes at 40 ° C, then 3.32 g (20 mmol) of 3,4-dimethoxybenzaldehyde is added and 10.8 ml (40 mmol) of tributylborate. The reaction mixture is stirred for 30 minutes at 50 ° C, after which 0.4 ml (5 mmol) of n-butylamine in 5 ml of anhydrous ethyl acetate are added dropwise over 15 minutes. The reaction mixture is heated to 70 ° C and stirred at this temperature for 10 hours, 30 ml of 1N aqueous hydrochloric acid are added and stirred at 60 ° C for another 2 hours, cooled, the organic layer is separated, and the aqueous layer is extracted with ethyl acetate (3 × 50 ml). The combined organic phases are dried over anhydrous sodium sulfate and concentrated in vacuo. The residue is recrystallized from methanol and dried. Receive: 1090 mg (55%) of the product.

Curcuminoid III Synthesis (PICUR)

To a solution of 350 mg (5.0 mmol) of boric acid anhydride in 60 ml of anhydrous ethyl acetate, 1.0 ml (10 mmol) of acetylacetone is added and the reaction mixture is stirred for 30 minutes at 40 ° C, after which 3.00 g (20 mmol) of heliotropin and 10.8 ml are added ( 40 mmol of tributylborate. The reaction mixture is stirred for 30 minutes at 50 ° C, after which 0.4 ml (5 mmol) of n-butylamine in 5 ml of anhydrous ethyl acetate are added dropwise over 15 minutes. The reaction mixture is heated to 70 ° C and stirred at this temperature for 10 hours, 30 ml of 1N aqueous hydrochloric acid are added and stirred at 60 ° C for another 2 hours, cooled, the organic layer is separated, and the aqueous layer is extracted with ethyl acetate (3 × 50 ml). The combined organic phases are dried over anhydrous sodium sulfate and concentrated in vacuo. The residue is recrystallized from methanol and dried. Receive: 673 mg (37%) of the product.

Synthesis of compounds of the invention

Synthesis of ethyl 4- (3,5-bis ((E) -3-hydroxy-4-methoxystyryl) -1H-pyrazol-1-yl) benzoate

To a solution of 368 mg (1 mmol) of Curcuminoid I in 10 ml of glacial acetic acid, 540 mg (3 mmol) of arylhydrazine are added as a solution in 2 ml of glacial acetic acid. The reaction mixture is heated with stirring to 65-70 ° C, 10 mg of anhydrous sodium acetate is added and maintained at a given temperature until the end of the reaction. Then, the reaction mixture is cooled and poured into 100 ml of ice-water, the precipitated precipitate is filtered off, dried and separated chromatographically by further recrystallizing the target fraction from dichloromethane / hexane. Receive: 307 mg (60%) of the product.

Synthesis of ethyl 4- (3,5-bis ((E) -3-hydroxy-4-methoxystyryl) -1H-pyrazol-1-yl) -2-fluorobenzoate

To a solution of 368 mg (1 mmol) of Curcuminoid I in 10 ml of glacial acetic acid, 594 mg (3 mmol) are added as a solution in 2 ml of glacial acetic acid. The reaction mixture is heated with stirring to 65-70 ° C and 10 mg of anhydrous sodium acetate is added and maintained at a given temperature until the end of the reaction. Then, the reaction mixture is cooled and poured into 100 ml of ice-water, the precipitated precipitate is filtered off, dried and separated chromatographically by further recrystallizing the target fraction from dichloromethane / hexane. Receive: 264 mg (50%) of the product.

Synthesis of ethyl 4- (3,5-bis ((E) -3-hydroxy-4-methoxystyryl) -1H-pyrazol-1-yl) benzoate

To a solution of 396 mg (1 mmol) of Curcuminoid I in 10 ml of glacial acetic acid, 540 mg (3 mmol) are added as a solution in 2 ml of glacial acetic acid. The reaction mixture is heated with stirring to 65-70 ° C, 1 drop of triethylamine is added and maintained at a given temperature until the end of the reaction. Then, the reaction mixture is cooled, poured into 100 ml of ice-water, the precipitated precipitate is filtered off, dried and separated chromatographically by further recrystallizing the target fraction from dichloromethane / hexane. 154 mg (34%) of the product are obtained.

Synthesis of 2,2,2-trifluoroethyl 4- (3,5-bis ((E) -4-hydroxy-3-methoxystyryl) -1H-pyrazol-1-yl) benzoate

Receiving 2,2,2-triptorelin-4-aminobenzoate

To a solution of 740 mg (3.01 mmol) of 2.2.2, -trifluoroethyl-4-nitrobenzoate (6) in 40 ml of ethanol, 1.32 ml (13 mmol) of cyclohexene and 200 mg of 10% palladium on carbon are added. The reaction mixture is boiled under argon for 15 hours, cooled, filtered through a pad of celite, the solvent is removed in vacuo, the residue is recrystallized from dichloromethane / hexane. Receive: 572 mg (87%) of the product.

Getting 2,2,2-triptorelin 4-hydrazinobenzoate

To a solution of 4.38 g (20 mmol) of p-aminobenzoic 2,2,2-trifluoroethyl ester in 30 ml of concentrated hydrochloric acid at -10 ° C, add 370 mg of sodium nitrite in 4 ml of water while maintaining the temperature below -10 ° C. The reaction mixture is stirred for 1.5 hours at a given temperature and a solution of 2.74 g (14.2 mmol) of tin (II) chloride dihydrate in 10 ml of concentrated hydrochloric acid is added, so that the temperature of the reaction mixture does not rise above -10 ° C and is stirred for another 2 hours at -10 ° C, and then 2 hours at room temperature. The precipitate is filtered off, dissolved in a minimal amount of hot water and 4N aqueous NaOH solution is added to pH 9. The resulting mixture is extracted with diethyl ether (3 × 50 ml), the organic phases are combined, washed with a saturated solution of sodium chloride (3 × 10 ml), the solvent is removed, the residue is dried in vacuo. Receive: 2.48 g (53%) technical product.

Synthesis of 2,2,2-trifluoroethyl 4- (3,5-bis ((E) -3-hydroxy-4-methoxystyryl) -1H-pyrazol-1-yl) benzoate

To a solution of 368 mg (1 mmol) of Curcuminoid I in 10 ml of glacial acetic acid, 702 mg (3 mmol) are added as a solution in 3 ml of glacial acetic acid. The reaction mixture is heated with stirring to 65-70 ° C, 1 drop of triethylamine is added and maintained at a given temperature until the end of the reaction. Then, the reaction mixture is cooled, poured into 100 ml of ice-water, the precipitated precipitate is filtered off, dried and separated by chromatography additionally by recrystallization of the target fraction from dichloromethane / hexane. Receive: 198 mg (35%) of the product.

Synthesis of ethyl 4- (3,5-bis ((E) -3-hydroxy-4-methoxystyryl) -1H-pyrazol-1-yl) -2-fluorobenzoate

To a solution of 396 mg (1 mmol) in 10 ml of glacial acetic acid, 594 mg (3 mmol) are added as a solution in 2 ml of glacial acetic acid. The reaction mixture is heated with stirring to 65-70 ° C, 1 drop of triethylamine is added and maintained at a given temperature until the end of the reaction. Then, the reaction mixture is cooled and poured into 100 ml of ice-water, the precipitated precipitate is filtered off, dried and separated chromatographically by further recrystallizing the target fraction from dichloromethane / hexane. 117 mg (25%) of the product are obtained.

Synthesis of ethyl 4- (3,5-bis ((E) -3,4-dimethoxystyryl) -1H-pyrazol-1-yl) -2-nitrobenzoate

Preparation of ethyl 4-hydrazino-2-nitrobenzoate

To a suspension of 4.20 g (20 mmol) of ethyl 4-hydrazino-2-nitrobenzoate in 30 ml of concentrated hydrochloric acid at -10 ° C add 370 mg of sodium nitrite in 4 ml of water, maintaining the temperature below -10 ° C. The reaction mixture is stirred for 1.5 hours at a given temperature, cooled to -35 ° C and, maintaining this temperature, a solution of 2.74 g (14.2 mmol) of tin (II) chloride dihydrate in 10 ml of concentrated hydrochloric acid is added, stirred for one hour at -30 ° C and then add 1N aqueous NaOH solution to pH 9. The resulting mixture is extracted with diethyl ether (3 × 50 ml), the organic phases are combined, washed with saturated sodium chloride solution (3 × 10 ml), the solvent is removed, the residue is dried in vacuum and separated by chromatography using ethyl acetate / di mixture Chloromethane 19: 1 Receive: 676 mg (15%) of the product.

Preparation of 4- (3,5-bis ((E) -3,4-dimethoxystyryl) -1H-pyrazol-1-yl) -2-nitrobenzoate

To a solution of 368 mg (1 mmol) of dimethylcurcuminoid I in 10 ml of glacial acetic acid, 675 mg (3 mmol) are added as a solution in 3 ml of glacial acetic acid. The reaction mixture is heated with stirring to 65-70 ° C, 1 drop of triethylamine is added and maintained at a given temperature until the end of the reaction. Then, the reaction mixture is cooled, poured into 100 ml of ice-water, the precipitated precipitate is filtered off, dried and separated by chromatography additionally by recrystallization of the target fraction from dichloromethane / hexane. Receive: 167 mg (30%) of the product.

Synthesis of 4- (3,5-bis ((E) -3,4-dimethoxystyryl) -1H-pyrazol-1-yl) -N-isopropylbenzamide

To a solution of 368 mg (1 mmol) in 10 ml of glacial acetic acid are added 675 (3 mmol) as a solution in 1 ml of glacial acetic acid. The reaction mixture is heated with stirring to 65-70 ° C, 1 drop of triethylamine is added and maintained at a given temperature until the end of the reaction. Then, the reaction mixture is cooled, poured into 100 ml of ice-water, the precipitated precipitate is filtered off, dried and separated by chromatography additionally by recrystallization of the target fraction from dichloromethane / hexane. Receive: 258 mg (44%) receiving.

Synthesis of 4- (3,5-bis ((E) -3,4-dimethoxystyryl) -1H-pyrazol-1-yl) -N-isobutylbenzamide

To a suspension of 512.5 mg (1 mmol) of acid in 5 ml of anhydrous DMF, 390 μl (2.2 mmol) of diisopropylethylamine are added, stirred for 30 minutes, and 570 mg (1.5 mmol) of HATU are added. The reaction mixture is stirred for 30 minutes at 40 ° C, 150 μl (1.5 mmol) of diisopropylethylamine are added and stirred at a given temperature for 12 hours, cooled, poured into 50 ml of water, the precipitate is filtered, washed on the filter with water, dried and separated by chromatography using as eluent, the system hexane: dichloromethane of increasing polarity. Receive: 380 mg (67%) of the product.

The use of chemical compounds according to the invention

Medication Use

The compounds described in this invention can be used to suppress multidrug resistance (MDR) in patients who receive various drugs for a long time. In particular, the compounds described in this invention are able to inhibit the protein of the ABC vector family P-glycoprotein. In addition, it has been shown that the use of a number of compounds constituting the present invention, together with cytostatic drugs (for example, doxorubicin), leads to an increase in the action of cytostatic drugs. The compounds described in this invention can also be used in the treatment of infectious diseases in which the pathogen has multidrug resistance (MDR), especially P-glycoprotein-mediated MDR, as examples of malaria (malaria plasmodia), tuberculosis, leishmaniasis and amebic dysentery. The administration of the compounds described in the present invention, together (separately, simultaneously or sequentially) with an agent for treating infectious diseases, is capable of potentiating the effect of the latter. The compounds described in this invention can also be used to increase the absorption or distribution of the drug. This method of administration includes the administration to the patient, separately, simultaneously or sequentially, of one of the compounds of the present invention and the aforementioned drug. In particular, this method can be used to increase the penetration of a drug into the central nervous system. For example, the compounds of the present invention can be used in a method for facilitating drug delivery across the blood-brain barrier.

Thus, we assume that the use of P-glycoprotein inhibitors, which are the subject of the present invention, in combination with current or new chemotherapy drugs against various oncological diseases or in combination with anti-infective drugs, for the treatment of infectious diseases, will achieve significant and long-term remission.

The method of therapeutic use of compounds

The subject matter of this invention also includes the administration to a subject in need of appropriate treatment of a therapeutically effective amount of a compound of the invention. A “therapeutically effective amount” is that amount of compound that is necessary to suppress multidrug resistance. In addition, the use of a therapeutically effective amount of a compound of the invention together with cytostatic drugs should lead to a detectable destruction of cancer cells or inhibition of their growth or rate of spread throughout the body, the size or number of tumors, or other characteristics of cancer. The exact amount required may vary from subject to subject, depending on the type, age and general condition of the patient, the severity of the disease, the characteristics of the anticancer agent, the method of administering the drug, the combined treatment with other drugs, etc. The compound, or a pharmaceutical composition containing the compound, may be administered to the patient in any amount and by any route of administration effective for suppressing multidrug resistance. The amount of a compound that will be effective in treating or preventing a particular disorder or condition depends, in particular, on well-known factors affecting the effective dosage of the drugs. In addition, in vitro or in vivo measurements may optionally be used to determine the optimal dose range. A rough way to determine the effective dose may be extrapolation of dose-response curves, which will depend on the model of in vitro or animal testing. The exact dosage level determined by the attending physician depends on well-known factors, including the method of administration of the drug, as well as the age, body weight, sex and general health of the patient; nature, severity and clinical condition of the disease; the use (or non-use) of concomitant therapy; and the nature and extent of genetic changes in the patient’s cells.

Pharmaceutically acceptable derivatives of the compounds

The compounds of this invention may exist in free form during processing, or, if required, in the form of a pharmaceutically acceptable salt or other derivative. The term "pharmaceutically acceptable salts" as used here refers to those salts that, within the framework of a medical report, are suitable for use in contact with human and animal tissues without undue toxicity, irritation, allergic reaction, etc., and meet a reasonable ratio of benefits. and risk. Examples of pharmaceutically acceptable non-toxic salts include salts formed with inorganic acids, such as hydrochloric, hydrobromic, phosphoric, sulfuric, and perchloric acids, or organic acids, such as acetic, oxalic, maleic, tartaric, succinic, citric, or malonic acids, or prepared by others methods used in this field, for example, using ion exchange. Other pharmaceutically acceptable ones include adipate and alfat heptanate, hexanate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate (mesylate), 2-naphthalenesulfonate, nicotinate, nitrate, oxalate, oxalate, pyentine sulfonate, nicotinate, nitrate, oxyate, oxalate, pyentine sulfonate, nicotinate, nitrate, oxalate, malsulfate (mesylate); tension, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate hemifumarate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate (tosylate), undekanat, valerate, and the like.

By "solvate" is meant the association or complex of one or more solvent molecules and a compound of the invention. Examples of solvents forming solvates include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine. The term "hydrate" refers to a complex where the solvent molecule is water.

Pharmaceutical compositions

The invention also relates to pharmaceutical compositions which contain at least one of the compounds described herein (or a prodrug form, a pharmaceutically acceptable salt or another pharmaceutically acceptable derivative) and one or more pharmaceutically acceptable carriers, solvents and / or excipients. These compositions may also contain one or more additional therapeutic agents. In addition, the compound of the present invention may be administered to a patient in need of appropriate therapy in combination with one or more other therapeutic agents.

The pharmaceutical compositions claimed in this invention contain the compounds of this invention together with pharmaceutically acceptable carriers, which may include any solvents, diluents, dispersions or suspensions, surfactants, isotonic agents, thickeners and emulsifiers, preservatives, astringents, lubricants materials, etc., suitable for a particular dosing form. Except for those cases where the medium of normal carriers is incompatible with the compound of the invention, for example, when any undesirable biological effects and other undesirable interactions occur with any other component (s) of the pharmaceutical composition, the use of such compositions is within the scope of this invention. Materials that can serve as pharmaceutically acceptable carriers include, but are not limited to, mono- and oligosaccharides, as well as their derivatives; malt, gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils such as peanut, cottonseed, saffrole, sesame, olive, corn and soybean oil; glycols such as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic solution, ringer's solution; ethyl alcohol and phosphate buffer solutions. Other non-toxic compatible lubricants, such as sodium lauryl sulfate and magnesium stearate, as well as dyes, release agents, film-forming agents, sweeteners, flavorings and flavoring agents, preservatives and antioxidants can also be included in the composition.

Dosage Forms

The subject of this invention are also dosage forms - a class of pharmaceutical compositions, the composition of which is optimized for a particular route of administration into the body in a therapeutically effective dose. The drug compositions of this invention can be administered orally, topically, rectally, intraocular, pulmonary, for example, as an inhalation spray, or intravascularly, intranasally, intraperitoneally, subcutaneously, intramuscularly, intrasternal, transdermal, and also by infusion, recommended dosages.

The dosage form of the present invention may contain a compound of the formula described herein, or a pharmaceutically acceptable salt, solvate or hydrate thereof, and any pharmaceutically acceptable carrier, adjuvant or solvent. The term "pharmaceutically acceptable carrier or adjuvant" means a carrier or adjuvant that can be introduced into a patient’s body together with a compound that is the essence of the present invention and does not destroy the pharmacological activity of this compound, and is non-toxic when administered in doses sufficient to deliver a therapeutic amount of compound.

Dosage forms of this invention may contain formulations obtained by the use of liposomes or microencapsulation methods, methods of preparing nanoform of the drug, and other examples known in the pharmaceutical industry.

Characterization of the biological activity of compounds

Study of the effect of the compounds of the invention on the accumulation of doxorubicin in cells with multidrug resistance

The biological activity of the compounds that are the subject of the present invention has been studied by various methods. For example, inhibition of P-glycoprotein by the accumulation of the substrate P-glycoprotein (doxorubicin) inside a tumor cell has been investigated. To study the accumulation of doxorubicin in cells with multidrug resistance and the effect of compounds that are the subject of the present invention on this process, cells of the K562 / Dox subline were scattered in 6-well plates. Doxorubicin (1 μM, 30 min.) Was added in the absence or presence of compounds (each at a concentration of 5 μM; the choice of concentration was determined by the range of toxic concentrations established in the MTT test). The cells were washed with cold physiological buffer, suspended in the same buffer on ice (to prevent doxorubicin leakage), and the fluorescence of the cells was analyzed. The fluorescence of the cells (hence, the accumulation of doxorubicin) was judged by the mean fluorescence channel in the FL1 or FL2, determined on the abscissa axis of the instrument. All experiments were performed in 2-3 replicates.

Illustrative examples of compounds (increasing the accumulation of doxorubicin by a factor of 1.2–10), which demonstrated activity against P-glycoprotein inhibition of the accumulation of the substrate P-glycoprotein (doxorubicin) inside a tumor cell:

Study of the ability of compounds according to the invention to inhibit P-glycoprotein

The effectiveness of P-glycoprotein inhibitors was investigated by determining the ability of compounds to enhance the cytotoxicity of doxorubicin (and other cytotoxic drugs) during joint incubation with resistant cells and the assessment of death by the colorimetric method. K562 / Dox subline cells were seeded into 96 well plates (Costar, United States) (104 cells in 190 μl of culture medium). Curcumin or compounds according to the invention were introduced into wells from serial dilutions of stock solutions. Served as control wells with a solvent (DMSO). The volume of the solution of the test compound did not exceed 5% of the volume of the culture medium in the wells. Each concentration of the drug was studied in 3 replications. Cells were incubated at 37 ° C, 5% CO ₂ for 72 hours. At the end of the incubation, 20 μl (50 μg) of the tetrazolium salt (3- (4,5-dimethylthiazol-2-yl) -5- (3-carboxymethoxyphenyl) -2- (4-sulfophenyl) -2P-tetrazolium, was introduced into the wells, salt), and the plates were placed in a CO ₂ incubator for 2 hours. Cell viability was judged by a color reaction that develops when tetrazolium salt is reduced to formazan by mitochondrial dehydrogenases. The color was recorded on a spectrophotometer (LKB, Sweden) at an excitation wavelength of 540 nm. The optical density in the wells, where the cells were incubated only with the solvent (control), was taken as 100%. The optical density in the wells with each concentration of the tested drugs were averaged and the percentage of surviving cells was calculated at a certain concentration of the studied drug.

IC ₅₀ = 1.4 μM IC ₅₀ = 0.21 μM

Examples of pharmaceutical compositions

The compounds described in this invention can be used for the prevention and treatment of human diseases, for example, in the form of the following compositions (in the examples below, the "Compound" means the active ingredient):

Tablet i mg / tablet Compound 50 Lactose Ph. Eur 223.75 Croscarmellose sodium 6.0 Corn starch 15 Polyvinylpyrolidone (5% vol. Pasta) 2.25 Magnesium stearate 3.0 Tablet II mg / tablet Compound 200 Lactose Ph. Eur 182.75 Croscarmellose sodium 12.0 Corn starch (5% vol. Paste) 2.25 Magnesium stearate 3.0 Capsule mg / capsule Compound ten Lactose Ph. Eur 488.5 Magnesia 1.5 The composition for injection I (with an active agent concentration of 50 mg / ml) Compound 5.0% vol. 1M sodium hydroxide solution 15.0% vol. 1M hydrochloric acid solution up to pH 7.6 Polyethylene glycol 400 4.5% vol. Water for injections up to 100% Aerosol I mg / ml Compound ten Sorbitan Trioleate 13.5 Trichlorofluoromethane 910.0 Dichlorodifluoromethane 490.0 Ointment ml Compound 40 mg Ethanol 0.3 ml Water 0.3 ml 1-dodecylazacycloheptanone 0.05 ml Propylene glycol up to 1 ml

These compositions can be prepared in accordance with standard pharmaceutical techniques. Tablets (I) - (II) can be enteric coated using, for example, cellulose acetate phthalate. Aerosol formulation (I) can be used in combination with standard dispensers; Sorbitan monooleate, sorbitan semi-oleate, polysorbate 80, polyglycerol oleate or oleic acid can be used as a suspending agent instead of sorbitan trioleate and soya lecithin.

Claims (30)

1. The compound of formula (I)

or its pharmaceutically acceptable salt, where: R ₁ , R ₂ , R ₃ , R ₄ - are chosen independently and represent H, halogen, NO ₂ ; R _x is selected from the group —C (= S) NHR ^A , —C (= O) OR ^A , —S (= O) ₂ NR ^A R ^B , —C (= O) NR ^A R ^B , 5-membered partially or fully unsaturated heterocycle containing 2 N atoms; R ^A - is chosen independently and represents a C _1-6 -alkyl, partially or fully halogenated C _1-8 -alkyl; R ^B - is chosen independently and represents H, C _1-8 -alkyl or partially or fully halogenated C _1-8 -alkyl; alternatively, in cases where any of R _x is -C (= O) NR ^A R ^B , R ^A and R ^B together with the nitrogen atom to which they are attached, can form a saturated cycle containing 5 atoms; R _p is independently selected and is C _1-6 alkyl, partially or fully halogenated C _1-6 alkyl; R _m is independently selected and is H, C _1-6 alkyl, partially or fully halogenated C _1-8 alkyl; alternatively, the adjacent R _p and R _m groups together with the two oxygen atoms to which they are attached, and the two carbon atoms of the benzene ring, can form a non-aromatic cycle containing from 5 to 6 atoms. 2. The compound according to claim 1, in which R ₁ , R ₂ , R ₃ , R ₄ - are chosen independently and represent H, halogen, NO ₂ ; R _x - represents -C (= S) NHR ^A , -C (= O) OR ^A , -S (= O) ₂ NR ^A R ^B , -C (= O) NR ^A R ^B, or a 5-membered partially or fully unsaturated heterocycle containing 2 N atoms; R ^A is independently selected and is C _1-6 alkyl, partially or fully halogenated C _1-8 alkyl; R ^B - is chosen independently and represents H, C _1-8 -alkyl, partially or fully halogenated C _1-8 -alkyl; alternatively, in cases where R _x is -C (= O) NR ^A R ^B , R ^A and R ^B together with the nitrogen atom to which they are attached, may form an unsaturated cycle containing 5 atoms; R _p - is chosen independently and represents CH ₃ , partially or fully halogenated C _1-6 -alkyl; R _m - is chosen independently and represents H, CH ₃ , partially or fully halogenated C _1-8 -alkyl. 3. The compound according to claim 1, selected from the group and representing:

4. The use of a compound according to claim 1 as an inhibitor of P-glycoprotein. 5. The use of a compound according to claim 1 for the preparation of a pharmaceutical composition for the treatment of diseases associated with the activity of P-glycoprotein. 6. The use according to claim 5, wherein the disease is characterized by multidrug resistance caused by the activity of P-glycoprotein. 7. Pharmaceutical composition for the treatment of diseases associated with the activity of P-glycoprotein, characterized in that it contains an effective amount of the compound according to claim 1 and a pharmaceutically acceptable carrier, solvent and / or excipient. 8. The pharmaceutical composition according to claim 7, which further comprises a cytostatic agent. 9. The pharmaceutical composition of claim 8, wherein the cytostatic agent is doxorubicin, vinblastine or methotrexate.