KR102625224B1 - Composition for preventing, improving or treating autoimmune diseases comprising pyrazol-one derivatives - Google Patents

Abstract

The present invention relates to a pharmaceutical composition for the prevention or treatment of autoimmune diseases comprising a pyrazol-one derivative, a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient. The compounds according to the present invention inhibit the activity of Peli1. It has the effect of effectively suppressing or inhibiting Peli1, so it can be used as a pharmaceutical composition that can fundamentally prevent or treat autoimmune diseases related to the activation of Peli1, as well as the development of biological agents for the treatment of autoimmune diseases and the development of Peli1 It can be used in various studies on the expression or inhibition of activity.

Description

Composition for preventing, improving or treating autoimmune diseases comprising pyrazol-one derivatives as an active ingredient {Composition for preventing, improving or treating autoimmune diseases comprising pyrazol-one derivatives}

The present invention relates to a composition containing a pyrazol-one derivative as an active ingredient, and more specifically, to a composition comprising a pyrazol-one derivative, a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient, for the prevention of autoimmune diseases, It relates to compositions for improvement or treatment.

Immunity is one of the body's self-protection systems against all foreign macromolecular substances (antigens) that invade or are injected into biological tissues. A major component of the immune system is lymphocytes, which are white blood cells produced in the bone marrow and circulating through the blood to lymphoid tissues and organs, mainly lymph nodes, spleen, and tonsils. As cells involved in the immune response, B cells proliferate rapidly when stimulated by an appropriate antigen, forming clones that produce special antibodies (immunoglobulins) to neutralize the antigen. The antibodies produced by B cells circulate in body fluids. While performing humoral immunity. Additionally, T cells are produced in the thymus and migrate to lymphoid tissue, and are responsible for cell-mediated immunity that directly attacks antigens.

Meanwhile, one of the most important characteristics of all normal individuals is that they do not react harmfully to antigens constituting the self, while they can recognize, react to, and eliminate many non-self antigens. It means having the ability. In this way, the unresponsiveness of a living body to self-antigens is called immunologic unresponsiveness or tolerance. If problems arise in inducing or maintaining this self-tolerance, an immune response occurs against self-antigens, which causes an attack on one's own tissues, leading to multiple sclerosis, type 1 diabetes, rheumatoid arthritis, Autoimmune diseases such as Hashimoto's thyroiditis may occur, and immune rejection reactions may occur after procedures such as transplantation.

Various methods are used to treat autoimmune diseases, and various methods are being tried to find more effective treatments. Treatment for most autoimmune diseases prescribes immunosuppressants that suppress excessive inflammatory responses rather than treatments that suppress the underlying cause of the disease. However, recently, through various studies, biological agents targeting IL-17 and IL-23, which were developed through research on T helper 17 (Th17) and Th1-related diseases, and biological agents targeting TNFa have been tested through clinical tests. The safety and effectiveness are being verified, and some are used in clinical trials for patients (Korean Patent Registration No. 10-1761506). However, although most of these agents are effective in relieving symptoms, they do not serve as a fundamental treatment for complete cure. In particular, there is still no known customized or targeted treatment for patients with severe psoriasis, rheumatoid arthritis, Sjögren's syndrome, multiple sclerosis, or inflammatory bowel disease. There is no bar.

The present inventors discovered compounds that inhibit the activity of the Peli1 (pellino homolog 1) gene, which is overexpressed and overactivated in autoimmune diseases, and confirmed that these compounds alleviate the symptoms of autoimmune diseases.

Accordingly, the purpose of the present invention is to provide a composition containing the above compounds that can prevent or treat autoimmune diseases such as psoriasis, inflammatory colitis, and rheumatoid arthritis.

In order to achieve the above object, the present invention provides a pharmaceutical composition for preventing or treating autoimmune diseases comprising a pyrazol-one derivative represented by the following formula (1), a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient. provides.

[Formula 1]

In Formula 1, R ¹ is hydrogen, C _1-5 alkyl, or -(CH ₂ ) _n COR ^a ,

Here, n is 1 to 3, R ^a is C _1-5 alkoxy,

R ² is C _1-5 alkyl or formula A below,

[Formula A]

where R ²¹ to R ²⁴ are independently hydrogen, halogen, C _1-5 alkyl, or C _1-5 alkoxy (* indicates the point of attachment of R ² to the nitrogen atom of Formula 1).

In addition, the present invention provides a pharmaceutical composition for preventing or treating autoimmune diseases comprising a pyrazol-one derivative represented by the following formula (2), a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient.

[Formula 2]

In Formula 2, R ³ is C _1-5 alkyl, C _1-3 haloalkyl, C _1-3 aminoalkyl, C _2-5 alkenyl, C _1-3 alkoxyalkyl or carbaldehyde, R ⁴ and R ⁵ are independently hydrogen, C _1-5 alkyl or hydroxylamino, and R ⁶ is the formula B below,

[Formula B]

where R ⁶¹ and R ⁶² are independently hydrogen, halogen, C _1-5 alkyl, or amine (* indicates the point of attachment of R ² to the nitrogen atom in Formula 2).

The pyrazol-one derivatives according to the present invention have the effect of effectively inhibiting or inhibiting the activity of Peli1 protein, and can be used as pharmaceutical compositions that can fundamentally prevent or treat autoimmune diseases related to activation of Peli1 protein. In addition, it can be used for various studies on the development of biological agents for the treatment of autoimmune diseases and the inhibition of the expression or activity of Peli1.

Figure 1 shows an in vitro autoubiquitination experiment using Ubc13, an E2 enzyme, to confirm the inhibitory effect of the compound according to the present invention on Peli1 protein activity, and immunoblotting using a Ub antibody. This shows the results of the analysis.
Figure 2 shows the results of confirming the expression of Peli1 protein in lesions of mouse models of psoriasis, rheumatoid arthritis (CIA), and inflammatory colitis (dextran sulfate sodium-induced colitis (DSS)) disease using immunochemical staining. will be.
Figure 3 shows the results of confirming the binding between Peli1 protein and IRF4 protein by performing Western blot.
Figure 4 shows the results of confirming by immunoblotting method whether the compound according to the present invention selectively inhibits ubiquitination of IRF4, a psoriasis-inducing downstream target protein by Peli1.
Figure 5 shows the results of confirming the efficacy of the compound according to the present invention in treating psoriasis through hematoxylin & eosin (H&E) staining.
Figure 6 shows the results of confirming the efficacy of the compound according to the present invention in treating psoriasis through hematoxylin & eosin (H&E) staining compared with the positive control group.
Figure 7 shows a comparison of the amount of change in skin thickness when treated with the compound according to the present invention and MTX, respectively.
Figure 8 shows a comparison of the change in weight loss between the group treated with the compound according to the present invention and the group treated only with DSS in an inflammatory colitis mouse model.
Figure 9 shows a comparison of the survival rates of the group treated with the compound according to the present invention and the group treated only with DSS in an inflammatory colitis mouse model.
Figure 10 shows a comparison of the disease activity index (DAI) of the group treated with the compound according to the present invention and the group treated only with DSS in an inflammatory colitis mouse model.
Figure 11 shows a comparison of the rate of bloody stool between the group treated with the compound according to the present invention and the group treated only with DSS in an inflammatory colitis mouse model.
Figure 12 shows a comparison of the structures of the colon tissues of the group treated with the compound according to the present invention and the group treated only with DSS in an inflammatory colitis mouse model by hematoxylin & eosin (H&E) staining.

The present invention provides a composition for preventing, improving, or treating autoimmune diseases, comprising a pyrazol-one derivative, a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient.

The present invention provides a pharmaceutical composition for preventing or treating autoimmune diseases comprising a pyrazol-one derivative represented by the following formula (1), a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient.

[Formula 1]

In Formula 1, R ¹ is hydrogen, C _1-5 alkyl, or -(CH ₂ ) _n COR ^a ,

Here, n is 1 to 3, R ^a is C _1-5 alkoxy,

R ² is C _1-5 alkyl or formula A below,

[Formula A]

where R ²¹ to R ²⁴ are independently hydrogen, halogen, C _1-5 alkyl, or C _1-5 alkoxy (* indicates the point of attachment of R ² to the nitrogen atom of Formula 1),

In one specific example, R ¹ is hydrogen, C _1-3 alkyl, or -(CH ₂ ) _n COR ^a , where n is 1 to 2, R ^a is C _1-3 alkoxy, and R ² is C _1-3 alkyl or the formula A′ below.

[Formula A`]

where R ²¹ to R ²⁴ are independently hydrogen, halogen, C _1-3 alkyl, or C _1-3 alkoxy (* indicates the point of attachment of R ² to the nitrogen atom of Formula 1),

In another specific example, R ¹ is hydrogen, methyl, ethyl or ethyloxycarbonyl (-CH ₂ COCH ₂ CH ₃ ), and R ²¹ to R ²⁴ are independently hydrogen, chlorine, bromine, methyl or It may be methoxy.

In another specific example, R ¹ may be hydrogen, methyl, ethyl, or ethyloxycarbonyl (-CH ₂ COOCH ₂ CH ₃ ), and R ² may be methyl, phenyl, 2-chlorophenyl, It may be 2,4-dichlorophenyl, 3,5-dichlorophenyl, 2-bromophenyl, 3-bromophenyl, m -tolyl, p -tolyl or 4-methoxyphenyl.

In another specific example, the compound of Formula 1 may be any one selected from the group consisting of the following compounds:

1) 4-(hydroxyimino)-2,5-dimethyl-4 H -pyrazol-3-one (4-(hydroxyimino)-2,5-dimethyl-4 H -pyrazol-3-one),

2) 4-(hydroxyimino)-5-methyl-2-( m -tolyl)-4 H -pyrazol-3-one (4-(hydroxyimino)-5-methyl-2-( m -tolyl)- 4H -pyrazol-3-one),

3) 2-(2-chlorophenyl)-4-(hydroxyimino)-5-methyl-4 H -pyrazol-3-one (2-(2-chlorophenyl)-4-(hydroxyimino)-5-methyl -4-pyrazol-3-one),

4) 2-(2,4-dichlorophenyl)-4-(hydroxyimino)-5-methyl-4 H -pyrazol-3-one (2-(2,4-dichlorophenyl)-4-(hydroxyimino) -5-methyl-4-pyrazol-3-one),

5) 2-(3,5-dichlorophenyl)-4-(hydroxyimino)-5-methyl-4 H -pyrazol-3-one (2-(3,5-dichlorophenyl)-4-(hydroxyimino) -5-methyl-4-pyrazol-3-one),

6) 2-(2-bromophenyl)-4-(hydroxyimino)-5-methyl-4 H -pyrazol-3-one (2-(2-bromophenyl)-4-(hydroxyimino)-5- methyl-4-pyrazol-3-one),

7) 2-(4-bromophenyl)-4-(hydroxyimino)-5-methyl-4 H -pyrazol-3-one (2-(4-bromophenyl)-4-(hydroxyimino)-5- methyl-4-pyrazol-3-one),

8) 4-(hydroxyimino)-5-methyl-2-( p -tolyl)-4 H -pyrazol-3-one (4-(hydroxyimino)-5-methyl-2-(p-tolyl)- 4-pyrazol-3-one),

9) 4-(hydroxyimino)-2-(4-methoxyphenyl)-5-methyl-4 H -pyrazol-3-one (4-(hydroxyimino)-2-(4-methoxyphenyl)-5- methyl-4-pyrazol-3-one),

10) 2-(2-chlorophenyl)-4-(methoxyimino)-5-methyl-4 H -pyrazol-3-one (2-(2-chlorophenyl)-4-(methoxyimino)-5-methyl -4-pyrazol-3-one),

11) 4-(methoxyimino)-5-methyl-2-( m -tolyl)-4 H -pyrazol-3-one (4-(methoxyimino)-5-methyl-2-( m -tolyl)- 4-pyrazol-3-one),

12) 4-(ethoxyimino)-5-methyl-2-phenyl-4 H -pyrazol-3-one (4-(ethoxyimino)-5-methyl-2-phenyl-4-pyrazol-3-one) , and

13) Ethyl 2-(((3-methyl-5-oxo-1-phenyl-1,5-dihydro-4 H -pyrazol-4-ylidene)amino)oxy)acetate (ethyl 2-(( (3-methyl-5-oxo-1-phenyl-1,5-dihydro-4 H -pyrazol-4-ylidene)amino)oxy)acetate.

In addition, the present invention provides a pharmaceutical composition for preventing or treating autoimmune diseases comprising a pyrazol-one derivative represented by the following formula (2), a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient.

[Formula 2]

In Formula 2, R ³ is C _1-5 alkyl, C _1-3 haloalkyl, C _1-5 aminoalkyl, C _2-5 alkenyl, C _{1- 3} alkoxyalkyl or carbaldehyde, R ⁴ and R ⁵ are independently hydrogen, C _1-5 alkyl or hydroxylamino, and R ⁶ is the formula B below,

[Formula B]

where R ⁶¹ and R ⁶² are independently hydrogen, halogen, C _1-5 alkyl, or amine (* indicates the point of attachment of R ² to the nitrogen atom in Formula 2).

As another specific example, in Formula 2, R ³ is methyl, ethyl, propyl, isopropyl, isobutyl, neopentyl, difluoromethyl, trifluoromethyl, chloromethyl, vinyl, aminomethyl, methyl, is toxymethyl or -COH, R ⁴ and R ⁵ are independently hydrogen, methyl, ethyl, isopropyl or -NHOH, and R ⁶ is chlorophenyl, dichlorophenyl, chlorofluorophenyl or aminochlorophenyl.

In another specific example, the compound of Formula 2 may be any one selected from the group consisting of the following compounds:

14) 2-(2-chlorophenyl)-5-methyl- 4H -pyrazol-3-one (2-(2-chlorophenyl)-5-methyl-4H - pyrazol-3-one),

15) 2-(3-chlorophenyl)-5-methyl-4 H -pyrazol-3-one (2-(3-chlorophenyl)-5-methyl-4 H -pyrazol-3-one),

16) 2-(4-chlorophenyl)-5-methyl-4 H -pyrazol-3-one (2-(4-chlorophenyl)-5-methyl-4 H- pyrazol-3-one),

17) 2-(2,3-dichlorophenyl)-5-methyl- 4H -pyrazol-3-one (2-(2,3-dichlorophenyl)-5-methyl-4H - pyrazol-3-one) ,

18) 2-(2,4-dichlorophenyl)-5-methyl-4 H -pyrazol-3-one (2-(2,4-dichlorophenyl)-5-methyl-4 H -pyrazol-3-one) ,

19) 2-(2,5-dichlorophenyl)-5-methyl- 4H -pyrazol-3-one (2-(2,5-dichlorophenyl)-5-methyl-4H - pyrazol-3-one) ,

20) 2-(2,6-dichlorophenyl)-5-methyl- 4H -pyrazol-3-one (2-(2,6-dichlorophenyl)-5-methyl-4H - pyrazol-3-one) ,

21) 2-(2-chloro-6-fluorophenyl)-5-methyl-4 H -pyrazol-3-one (2-(2-chloro-6-fluorophenyl)-5-methyl-4 H -pyrazol -3-one),

22) 2-(2-amino-2-chlorophenyl)-5-methyl-4 H -pyrazol-3-one 2-(4-amino-2-chlorophenyl)-5-methyl-4 H -pyrazol-3 -one,

23) 2-(2-chlorophenyl)-5-ethyl-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-ethyl-4 H -pyrazol-3-one),

24) 2-(3-chlorophenyl)-5-ethyl-4 H -pyrazol-3-one (2-(3-chlorophenyl)-5-ethyl-4 H -pyrazol-3-one),

25) 2-(4-chlorophenyl)-5-ethyl-4 H -pyrazol-3-one (2-(4-chlorophenyl)-5-ethyl-4 H -pyrazol-3-one),

26) 2-(2,4-dichlorophenyl)-5-ethyl-4 H -pyrazol-3 - one),

27) 2-(2-chloro-6-fluorophenyl)-5-ethyl-4 H -pyrazol-3-one (2-(2-chloro-6-fluorophenyl)-5-ethyl-4 H -pyrazol -3-one),

28) 2-(2-chlorophenyl)-5-propyl-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-propyl-4 H -pyrazol-3-one),

29) 2-(3-chlorophenyl)-5-propyl-4 H -pyrazol-3-one (2-(3-chlorophenyl)-5-propyl-4 H -pyrazol-3-one),

30) 2-(4-chlorophenyl)-5-propyl-4 H -pyrazol-3-one (2-(4-chlorophenyl)-5-propyl-4 H -pyrazol-3-one),

31) 2-(2,4-dichlorophenyl)-5-propyl- 4H -pyrazol-3- one ) ,

32) 2-(2,5-dichlorophenyl)-5-propyl-4 H -pyrazol-3-one (2-(2,5-dichlorophenyl)-5-propyl-4 H -pyrazol-3-one) ,

33) 2-(2-chloro-6-fluorophenyl)-5-propyl-4 H -pyrazol-3-one (2-(2-chloro-6-fluorophenyl)-5-propyl-4 H -pyrazol -3-one),

34) 2-(2 - chlorophenyl)-5-isopropyl-4 H -pyrazol-3-one),

35) 2-(2-chlorophenyl)-5-isobutyl-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-isobutyl-4 H -pyrazol-3-one),

36) 5-(tert-butyl)-2-(2-chlorophenyl)-4 H -pyrazol-3-one (5-(tert-butyl)-2-(2-chlorophenyl)-4 H -pyrazol- 3-one),

37) 2-(2-chlorophenyl)-5-(difluoromethyl)-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-(difluoromethyl)-4 H -pyrazol-3 -one),

38) 2-(2-chlorophenyl)-5-(trifluoromethyl)-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-(trifluoromethyl)-4 H -pyrazol-3 -one),

39) 2-(2-chlorophenyl)-5-(chloromethyl)-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-(chlororomethyl)-4 H -pyrazol-3-one ),

40) 2-(4-chlorophenyl)-5-vinyl-4 H -pyrazol - 3-one),

41) 5-(aminomethyl)-2-(2-chlorophenyl)-4 H -pyrazol-3-one (5-(aminomethyl)-2-(2-chlorophenyl)-4 H -pyrazol-3-one ),

42) 2-(2-chlorophenyl)-5-(methoxymethyl)-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-(methoxymethyl)-4 H -pyrazol-3- one),

43) 2-(2-chlorophenyl)-4,5-dimethyl- 4H -pyrazol-3-one (2-(2-chlorophenyl)-4,5-dimethyl-4H - pyrazol-3-one) ,

44) 2-(2-chlorophenyl)-4-ethyl-5-methyl-4 H -pyrazol-3-one (2-(2-chlorophenyl)-4ethyl-5-methyl-4 H -pyrazol-3- one),

45) 2-(2-chlorophenyl)-5-ethyl-4-methyl-4 H -pyrazol-3-one (2-(2-chlorophenyl)-5-ethyl-4-methyl-4 H -pyrazol- 3-one),

46) 2-(2-chlorophenyl)-4-isopropyl-5-(trifluoromethyl)-4 H -pyrazol-3-one (2-(2-chlorophenyl)-4-isopropyl-5-methyl -4H -pyrazol-3-one),

47) 2-(2-chlorophenyl)-4-ethyl-5-(trifluoromethyl)-4 H -pyrazol-3-one (2-(2-chlorophenyl)-4-methyl-5-(trifluoromethyl )-4 H -pyrazol-3-one),

48) 2-(2-chlorophenyl)-4-(hydroxyamino)-4,5-dimethyl-4 H -pyrazol-3-one 2-(2-chlorophenyl)-4-(hydroxyamino)-4, 5-dimethyl- 4H -pyrazol-3-one, and

49) 1-(2-chlorophenyl)-5-oxo-4,5-dihydro-1 H -pyrazole-3-carbaldehyde (1-(2-chlorophenyl)-5-oxo-4,5-dihydro -1H -pyrazole-3-carbaldehyde).

The term "pyrazol-one derivative" of the present invention is chemically 4-(hydroxyimino)-5-methyl-4 H -pyrazol-3-one (4-(hydroxyimino)-5-methyl-4 H - pyrazol-3-one) or 4 H -pyrazol-3-one (4 H -pyrazol-3-one). The pyrazol-one derivative inhibits the expression of Peli1 and can prevent or treat diseases related to its activation, especially autoimmune diseases.

The term "Pelli 1 (pellino homolog 1) gene" of the present invention refers to a gene encoding the pellino homolog 1 protein, known as an E3 ubiquitin conjugation enzyme, and is present on chromosome 2 in humans and chromosome 11 in mice. It exists in chromosomes. The specific base sequence and protein information of the protein or gene are known in NCBI (NCBI Reference Sequence: NP_065702, NM_020651).

The term "pharmaceutically acceptable salt" or "foodologically acceptable salt" of the present invention refers to a salt that can be used pharmaceutically among salts that are substances in which cations and anions are bound by electrostatic attraction. Typically, it can be a metal salt, a salt with an organic base, a salt with an inorganic acid, a salt with an organic acid, a salt with a basic or acidic amino acid, etc. For example, the metal salt may be an alkali metal salt (sodium salt, potassium salt, etc.), an alkaline earth metal salt (calcium salt, magnesium salt, barium salt, etc.), an aluminum salt, etc. Salts with bases include triethylamine, pyridine, picoline, 2,6-lutidine, ethanolamine, diethanolamine, triethanolamine, cyclohexylamine, dicyclohexylamine, N,N -dibenzylethylenediamine, etc. It can be a salt of . Salts with inorganic acids can be salts with hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid, etc. Salts with organic acids may include salts with formic acid, acetic acid, trifluoroacetic acid, phthalic acid, fumaric acid, oxalic acid, tartaric acid, maleic acid, citric acid, succinic acid, methanesulfonic acid, benzenesulfonic acid, and p -toluenesulfonic acid. Salts with basic amino acids can be salts with arginine, lysine, ornithine, etc. Salts with acidic amino acids can be salts with aspartic acid, glutamic acid, etc.

For the purpose of the present invention, the pharmaceutically acceptable salt or foodologically acceptable salt is an acid addition salt or base of a pyrazol-one derivative suitable for the treatment of patients expected to develop an autoimmune disease or suffering from the disease. It may be interpreted as an addition salt, but is not particularly limited thereto.

The term "autoimmune disease" of the present invention refers to a group consisting of psoriasis, rheumatoid arthritis, multiple sclerosis, lupus, Bachet's disease, ankylosing spondylitis, Sjögren's syndrome, Crohn's disease, and inflammatory bowel disease, which are caused by the expression of Peli1 protein. It may be any one or more selected from.

The term "prevention" of the present invention refers to administering a pharmaceutical composition containing a pyrazol-one derivative, a pharmaceutically acceptable salt thereof, or an isomer thereof provided by the present invention as an active ingredient to an individual expected to develop an autoimmune disease. This means all actions that suppress or delay the onset of the above disease.

The terms “improvement” and “treatment” in the present invention refer to any clinical intervention to alter the natural processes of the individual or cell to be treated, performed during the course of a clinical pathology or to prevent it. can do. The desired therapeutic effects include preventing the occurrence or recurrence of the disease, alleviating symptoms, reducing all direct or indirect pathological consequences of the disease, preventing metastasis, reducing the rate of disease progression, and alleviating the disease state. Or it includes temporary relief, remission, or improvement of prognosis. For the purpose of the present invention, the treatment refers to the course of the autoimmune disease by administering a pharmaceutical composition containing a pyrazol-one derivative, a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient to a patient suffering from an autoimmune disease. It can be interpreted to include all acts that cause aggression, but is not particularly limited thereto.

In the present invention, “C _1-5 alkyl” refers to a linear or branched saturated hydrocarbon group having 1 to 5 carbon atoms. For example, but not limited to, methyl, ethyl, propyl, butyl, pentyl, 1-methylethyl, 1-methylpropyl, 2-methylpropyl, 1,1-dimethylethyl, 1-methylbutyl or 1, It is 1-dimethylpropyl.

In the present invention, “C _1-3 alkyl” refers to a linear or branched saturated hydrocarbon group having 1 to 3 carbon atoms. For example, but not limited to, methyl, ethyl, propyl or isopropyl.

In the present invention, “C _1-3 haloalkyl” refers to a functional group in which one or more hydrogens are replaced with halogen in a linear or branched saturated hydrocarbon group having 1 to 3 carbon atoms. For example, but not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, chloroethyl or chloropropyl.

In the present invention, “C _1-5 aminoalkyl” means a functional group in which hydrogen or an alkyl group is bonded to a nitrogen atom and a saturated hydrocarbon group having 3 to 5 carbon atoms. Examples, but not limited thereto, include aminomethyl, dimethylaminomethyl, methylethylaminomethyl, diethylaminomethyl, aminoethyl, dimethylaminoethyl, methylethylaminoethyl, diethylaminoethyl, aminopropyl, dimethylaminopropyl. , methylethylaminopropyl, diethylaminopropyl or 1-(dibutylamino)propyl.

In the present invention, “C _2-5 alkenyl” refers to an unsaturated hydrocarbon group having 2 to 5 carbon atoms.

In the present invention, “C _1-5 alkoxy” is an OR group where R is a C _1-5 alkyl group as defined above. For example, but not limited to, methoxy, ethoxy, propoxy, butoxy, pentoxy, 1-methylethoxy, 1-methylpropoxy, 1-methylbutoxy, 1,1-dimethyl. Toxy, t- butoxy.

In the present invention, “C _1-3 alkoxyalkyl” is a functional group in which the alkoxy group is bonded to a C _1-5 alkyl group as defined above. Examples, but not limited to, are methoxymethyl, ethoxyethyl, propoxymethyl, butoxypropyl or pentoxybutyl.

In the present invention, “halogen” is a fluorine, chlorine, bromine or iodine atom.

Meanwhile, the pharmaceutical composition of the present invention may additionally contain appropriate carriers, excipients, or diluents commonly used in the preparation of pharmaceutical compositions. The composition containing a pharmaceutically acceptable carrier may be in various oral or parenteral dosage forms. When formulated, it can be prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants. Solid preparations for oral administration may include tablets, powders, granules, capsules, etc. These solid preparations include one or more compounds and at least one excipient, such as starch, calcium carbonate, sucrose, or lactose. It can be prepared by mixing (lactose), gelatin, etc.

Additionally, in addition to simple excipients, lubricants such as magnesium stearate, talc, etc. may also be used. Liquid preparations for oral administration include suspensions, oral solutions, emulsions, and syrups. In addition to the commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. there is. Preparations for parenteral administration may include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable esters such as ethyl oleate. As a base for suppositories, witepsol, macrogol, tween 61, cacao, laurel, glycerogelatin, etc. can be used.

In addition, the pharmaceutical composition of the present invention includes tablets, pills, powders, granules, capsules, suspensions, oral solutions, emulsions, syrups, sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations and suppositories. It may have any one formulation selected from.

Through examples, the present inventors experimentally confirmed that the pyrazol-one derivatives of the present invention, their pharmaceutically acceptable salts, or isomers thereof are effective in preventing or treating autoimmune diseases.

In one example of the present invention, it was confirmed that the compound according to the present invention specifically reacts with Peli1 protein and has the effect of inhibiting the activity of Peli1 protein (see Example 1).

In another example of the present invention, it was confirmed that the expression of Peli1 was significantly increased in autoimmune diseases (psoriasis, rheumatoid arthritis, inflammatory colitis) (see Example 2).

In another embodiment of the present invention, the expression of IRF4 protein is abnormally increased by Peli1 protein, resulting in the development of autoimmune disease, and the compound according to the present invention selectively inhibits ubiquitination of IRF4, a psoriasis-causing downstream target protein, by Peli1. It was confirmed that autoimmune diseases can be treated by inhibiting (see Example 3).

In another example of the present invention, the psoriasis treatment effect of the compound according to the present invention was confirmed using a BALB/c mouse model treated with the psoriasis-inducing substance imiquimod (IMQ) (see Example 5).

In another example of the present invention, the inflammatory colitis treatment effect of the compound according to the present invention was confirmed using a DSS-treated inflammatory enteritis mouse model (see Example 6).

As another embodiment, the present invention provides a method for preventing or treating autoimmune diseases comprising administering to an individual a composition containing a pyrazol-one derivative, a pharmaceutically acceptable salt thereof, or an isomer thereof as an active ingredient. provides.

The term “individual” in the present invention refers to all animals, including humans, that are likely to develop or have the autoimmune disease. By administering the composition of the present invention to an individual, autoimmune diseases can be alleviated or treated.

In the present invention, the term "alleviation" refers to all actions in which an autoimmune disease is improved or beneficial by administration of the composition according to the present invention.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount.

In the present invention, the term "administration" refers to introducing the pharmaceutical composition of the present invention into a subject by any appropriate method, and the administration route can be administered through various routes such as oral or parenteral as long as it can reach the target tissue. .

The pharmaceutical composition of the present invention can be appropriately administered to an individual according to the conventional method, administration route, and dosage used in the art, depending on the purpose or need. Examples of routes of administration include oral, parenteral, subcutaneous, intraperitoneal, intrapulmonary, and intranasal administration; parenteral infusion includes intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration.

In addition, the appropriate dosage and number of administrations can be selected according to methods known in the art, and the amount and number of administrations of the pharmaceutical composition of the present invention actually administered are determined by the type of symptom to be treated, administration route, gender, and health condition. , can be appropriately determined by various factors such as diet, age and weight of the individual, and severity of the disease.

As used herein, the term "pharmaceutically effective amount" refers to an amount sufficient to suppress or relieve pain in chemotherapy-induced peripheral neuropathy with a reasonable benefit/risk ratio applicable to medical use, and the effective dose level is determined by the type of individual. and severity, age, gender, activity of the drug, sensitivity to the drug, time of administration, route of administration and excretion rate, duration of treatment, factors including concurrently used drugs, and other factors well known in the medical field. The composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. And it can be administered single or multiple times. Considering all of the above factors, it is important to administer an amount that can achieve maximum effect with the minimum amount without side effects, and can be easily determined by a person skilled in the art.

Meanwhile, the composition of the present invention can also be used as a food composition to prevent or improve autoimmune diseases.

When the composition of the present invention is used as a food composition, in addition to the pyrazol-one derivative, its foodologically acceptable salt, or isomer thereof as the active ingredient, components commonly added during food production, such as proteins, carbohydrates, It may additionally contain fats, nutrients, seasonings and flavoring agents.

Examples of such carbohydrates include monosaccharides (eg, glucose, fructose, etc.); Common sugars such as disaccharides (e.g. maltose, sucrose, oligosaccharides, etc.) and polysaccharides (e.g. dextrin, cyclodextrin, etc.) and sugar alcohols such as xylitol, sorbitol, and erythritol. As a flavoring agent, natural flavoring agents such as thaumatin and stevia extract (e.g., rebaudioside A, glycyrrhizin) and synthetic flavoring agents such as saccharin and aspartame can be used.

There are no particular restrictions on the type of food. Examples of foods to which the food composition of the present invention can be added include meat, sausages, bread, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, and tea. , drinks, alcoholic beverages, and vitamin complexes, etc., and includes all health foods in the conventional sense.

When the food composition of the present invention is manufactured as a drink, citric acid, high fructose corn syrup, sugar, glucose, acetic acid, malic acid, fruit juice, etc. are added as active ingredients in addition to pyrazol-one derivatives, food-acceptable salts thereof, or isomers thereof. Can be included.

In addition to the above-mentioned ingredients, the food composition contains various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH regulators, stabilizers, preservatives, glycerin, and alcohol. , may contain carbonating agents used in carbonated beverages, etc. In addition, the food composition of the present invention may contain pulp for the production of natural fruit juice, beverages, and vegetable beverages. These ingredients can be used independently or in combination.

Below, preferred embodiments are presented to aid understanding of the present invention. However, the following examples are provided only to make the present invention easier to understand, and the content of the present invention is not limited by the following examples.

[Example]

Example 1. Analysis of the effect of the compound according to the present invention on inhibiting Peli1 protein activity

1-1. Confirmation of activity inhibition effect of Peli1

To confirm the inhibitory effect of the compound according to the present invention on Peli1 protein activity, an in vitro autoubiquitination experiment was performed on compound 10 and its isomers using Ubc13, an E2 enzyme, and immunoblotting using a Ub antibody. Analysis was performed by immunoblotting. The major isomer of compound 10 is a cis-type isomer, and the minor isomer is a trans-type. Experiments were conducted using isomer ratios of cis and trans at 2:1 (mix) and 4:1 (ori), or using only the cis form (maj) in three different compositions.

As a result, as shown in Figure 1, all of these compounds were found to inhibit the activity of Peli1 protein, and it was confirmed that among them, the 2:1 mixture most effectively inhibited the activity of Peli1 protein.

1-2. Verification of specific reaction with Peli1

The specificity (selectivity) for Peli1 was verified by confirming whether the compound according to the present invention inhibits the activation of other types of E3 ubiquitin conjugating enzymes in addition to the Peli1 protein.

In order to confirm whether the pyrazol-one derivative according to the present invention selectively inhibits the activity of Peli1 protein, eight different types of E3 ubiquitin conjugating enzymes were isolated and purified and analyzed to see whether they inhibited autoubiquitination. After analysis by immunoblotting using a Ub antibody, the E2 enzyme used for each E3 ubiquitin conjugation enzyme by compound 10 and the degree of inhibition of autoubiquitination thereof were relatively quantified. The results are shown in Table 2 below. Each E3 conjugation enzyme was fused with GST through separation and purification. As a result, the compound according to the present invention did not show an inhibitory reaction on most other E3 conjugation enzymes other than Peli1 protein.

Example 2. Confirmation of Peli1 protein expression level in autoimmune disease

By confirming the expression of Peli1 protein in lesion tissues of Th17 T helper-related diseases such as psoriasis, rheumatoid arthritis, and inflammatory colitis animal model (Peli1-induced expression mouse) using immunostaining, the expression level of peli1 protein in autoimmune diseases was confirmed. Confirmed.

Psoriasis was induced in the animal model by periodically treating the skin with imiquimod cream (total of 7 treatments), and collagen and adjuvant were administered subcutaneously to the tail to induce collagen-induced arthritis. arthritis, CIA) was induced. Additionally, DSS (dextran sodium sulfate) was dissolved in drinking water and treated for about 7 to 10 days to induce inflammatory colitis (dextran sulfate sodium-induced colitis). The tissues of the animal model that induced psoriasis, rheumatoid arthritis, and inflammatory colitis and the animal model (control) not treated with these substances were confirmed using immunostaining.

As a result, as shown in Figure 2, it was confirmed that the expression of Peli1 was significantly increased in the epidermal layer of the skin of the psoriasis animal model, and the expression of Peli1 was also confirmed in the immune cells infiltrated in the dermal layer. In addition, the expression of Peli1 was confirmed to be increased in overproliferated synovial cells (synoviocytes) of joint disease in an animal model of rheumatoid arthritis, and in colonic lesions of an inflammatory colitis animal model, the expression of Peli1 was increased in macrophage cells infiltrating the lesion. It was confirmed that there was a significant increase.

Example 3. Confirmation of the ubiquitination inhibitory effect of IRF4 protein, a binding protein of Peil1 protein

3-1. Confirmation of binding of Peil1 protein and IRF4 protein

During the development of psoriasis, the expression of IRF4 protein is abnormally increased by Peli1 protein. Therefore, to confirm the binding between Peli1 protein and IRF4 protein, lysate of human keratinocytes cell (HaCaT) was incubated with anti-Peli1, and then Peli1 protein and IRF4 protein were incubated using anti-IRF4. The binding between the two was confirmed by performing western blot.

As a result, as shown in Figure 3, it was confirmed that Peli1 protein and IRF4 protein bind.

3-2. Confirmation of the IRF4 ubiquitination inhibition effect of the compound according to the present invention

To determine whether the compound according to the present invention (compound 10) selectively inhibits ubiquitination of IRF4, a psoriasis-causing downstream target protein by Peli1, Myc-Peli1, HA-IRF4, and Ub expression vectors were transfected into HaCaT cell line. Afterwards, immunoprecipitation was performed on these lysates with an HA antibody, and an immunoblotting experiment was performed with a Ub antibody.

As a result, as shown in Figure 4, it was confirmed that compound 10 very strongly inhibited in vivo ubiquitination of IRF4, a target protein by Peli1. In FIG. 4, #1 and #3 represent low-molecular-weight compounds of the positive control, and FL represents Peli1 full length (wild-type).

Example 4. Confirmation of psoriasis treatment effect

The psoriasis treatment effect of Compound 10 according to the present invention was confirmed using a BALB/c mouse model treated with the psoriasis-inducing substance imiquimod (IMQ).

The BALB/c mouse model was first treated with the psoriasis-inducing substance IMQ for 3 days, and on the 4th day, it was treated with vaseline and IMQ, and simultaneously with the control vehicle (vehicle, 5% DMSO + 5% cremorphol EL + 90% PBS). The compound according to the invention (compound 10, compound dissolved in 5% DMSO + 5% cremorphol EL + 90% PBS) (100 mg/kg) was injected intraperitoneally until the 7th day. Afterwards, the skin of the mouse model was sampled and the psoriasis treatment effect was confirmed through hematoxylin & eosin (H&E) staining. As a result, as shown in Figure 5, when treated with Compound 10, which inhibits Peli1 activity, it was observed that skin thickness decreased and red spots were alleviated compared to the control group. From this, it can be seen that the compound according to the present invention is effective in treating psoriasis.

Meanwhile, under experimental conditions similar to the above, methotrexate (MTX, 1 mg/kg), which is commonly prescribed clinically as a treatment for psoriasis, was intraperitoneally injected into mice every day for 8 days simultaneously with IMQ and applied to the back, and skin tissue was stained with H&E. The psoriasis treatment effect was analyzed (positive control group). As a result, as shown in Figure 6, it was confirmed that when treated with MTX, which was used as a positive control, the thickness of the skin decreased compared to when treated with only PBS.

Afterwards, in order to compare the psoriasis treatment efficacy in the above experiments, skin thickness was measured and quantified. As a result, as shown in Figure 7, when treated with MTX, skin thickness was found to be reduced by 40%, and when treated with Compound 10 (IMQ 100), skin thickness was shown to be reduced by 45%. Through this, it was confirmed that the psoriasis treatment efficacy of Compound 10, a pyrazol-one derivative according to the present invention, is higher than that of MTX.

Example 5. Confirmation of inflammatory colitis treatment effect

When dextran sulfate sodium (DSS) is dissolved in distilled water and administered, the intestinal epithelium layer of mice collapses and various types of immune cells present in the laminar propria are destroyed. When exposed to the outside, immune cells are activated by antigens present in the intestines and induce an excessive inflammatory response, leading to inflammatory colitis.

In this example, in order to confirm the effectiveness of the compound according to the present invention in treating inflammatory colitis, DSS was dissolved in drinking water at a concentration of 3% and administered to a mouse model for a week, while Compound 10 was administered intraperitoneally at a concentration of 100 mg/kg.

As a result, as shown in Figures 8 and 9, as a result of analyzing the changes in weight loss and survival rate of the inflammatory colitis mouse model, rapid weight loss and decreased survival rate were observed in the DSS-treated experimental group (DSS-Vehicle). On the other hand, the experimental group treated with Compound 10 (DSS-Compound 11) showed little change in weight loss and survival rate despite treatment with DSS, and most of the experimental group treated with DSS died 7 days after intraperitoneal injection. Unlike (DSS-Vehicle), it was confirmed that the mortality rate was extremely low.

In addition, the wateriness and color of stool, which are standard indicators of inflammatory colitis, were scored and combined with the weight loss ratio score to create a disease activity index (DAI), so that the group treated with DSS alone and the group treated with DSS and Compound 10 together. was compared.

As a result, as shown in Figure 10, from 5 days of DSS treatment, the group treated only with DSS (DSS-Vehicle) showed a continuous significant increase in DAI, but the group treated with Compound 10 and DSS together (DSS-Compound) 10) confirmed that the DAI score decreased from the 7th day.

In addition, as shown in Figure 11, all mice in the group treated only with DSS (DSS-V) showed bloody stool on day 7, while only 25% of the mice treated with Compound 10 (DSS-Compound 10) showed bloody stool on day 7. .

Meanwhile, in order to confirm the therapeutic efficacy of inflammatory colitis according to the treatment of the compound according to the present invention, the colon tissues of the above experimental groups were subjected to hematoxylin & eosin (H&E) staining and their structures were compared.

As a result, as shown in Figure 12, the structure of the intestinal epithelium of the DSS-only treated group (DSS Vehicle) was confirmed to be damaged compared to the control vehicle (Control Vehicle). On the other hand, in the group treated with Compound 10 (DSS Compound 10), it was confirmed that the intestinal epithelial structure was not as perfect as the control group, but maintained a close shape.

From this, it can be seen that the compound according to the present invention shows a very excellent effect in the treatment of inflammatory colitis.

From the results of the above examples, it was found that pyrazol-one, a pharmaceutically acceptable salt thereof, or an isomer thereof according to the present invention can prevent or treat autoimmune diseases expressed by the Peli1 protein.

The description of the present invention stated above is for illustrative purposes, and a person skilled in the art to which the present invention pertains can understand that it can be easily modified into other specific forms without changing the technical idea or essential features of the present invention. There will be. Therefore, the embodiments described above should be understood in all respects as illustrative and not restrictive.

Claims (10)

A pharmaceutical composition for preventing or treating autoimmune diseases comprising a pyrazol-one derivative represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient:
[Formula 1]

In Formula 1, R ¹ is methoxy,
R ² is 2-chlorophenyl,
The autoimmune disease is characterized by psoriasis or inflammatory colitis. delete delete delete delete delete delete delete According to paragraph 1,
The pyrazol-one derivative is a pharmaceutical composition for preventing or treating autoimmune diseases, characterized in that it inhibits the expression of Peli1 protein. delete