WO2013107164A1 - Novel indoleamine-2, 3-dioxygenase inhibitor, preparation method therefor and uses thereof - Google Patents

Abstract

The present invention relates to a novel indoleamine-2, 3-dioxygenase (IDO) inhibitor, a preparation method therefor, and the uses thereof. The inhibitor is a compound as represented by formula (I) or a salt, a solvate, a polymorph, an enantiomer, or a racemic mixture pharmaceutically acceptable thereto. R¹ or R² is independently selected from hydrogen, a C₁-C₅ alkyl group, a halogen, and a substituent group as represented by formula (II), and one of the R¹ and the R² is the substituent group as represented by formula (II). R³ is selected from an aryl group, and n=0, 1, 2, or 3. Compared with a known IDO inhibitor, the novel inhibitor has a more powerful IDO inhibitory effect, and can be used to treat diseases having a pathological feature of an IDO-mediated tryptophan metabolic pathway, for example, tumors, cancers, Alzheimer disease, autoimmune diseases, cataract, mental disorders, depression, and/or anxiety disorders. The preparation method therefor has the advantages of easy operation and a mild reaction condition, and saves the solvent and reduces pollution, thereby facilitating industrial production.

Description

 Novel guanamine- 2,3-hanoxygenase inhibitor, preparation method and use thereof

 The invention belongs to the field of medicinal chemistry, and particularly relates to a tryptophan guanamine- 2,3-dioxygenase inhibitor containing a 1,2,3-triazole structure, a preparation method thereof and use thereof. Background technique

 Indole-2,3-dioxygenase (IDO; MW 48,000; EC 1.13.11.42) is a heme-containing enzyme that is the first enzyme in the mammalian tryptophan metabolism pathway and is Rate limiting enzyme. IDO catalyzes the oxidation of the essential amino acid monotryptophan by the conversion of hydrogen to N-nonanoyl kynurenine and is responsible for the cleaning of tryptophan in the human body. By degrading tryptophan, IDO causes a microenvironment in which tryptophan is absent in the body, which in turn leads to the development of diseases such as cancer, cataracts, and neurological disorders that are closely related to the loss of tryptophan. Therefore, the search for high-efficiency inhibitors based on IDO targets has become a research hotspot in drug development in recent years. It is currently known that certain tryptophan (substrate) analogs such as 1-mercapto-L-tryptophan (1-MT) and β-(3-benzofuranyl)-DL-alanine are IDO Competitive inhibitors (Cady, SG and Sono, M. Arch. Biochem. Biophys. 1991, 291, 326).

 Interferon Y is one of several potential IDO expression inducers. During sustained activation of high levels of interferon gamma stimulation, IDO reduces the availability of free serum tryptophan and thus reduces the production of serotonin. These changes, combined with the accumulation of neuroactive kynurenine metabolites such as quinolinic acid (also induced by IDO), promote the development of neuropathy/psychiatric disorders and are a cause of multiple psychological disorders, as well as IDO activation and Tryptophan degradation is a cause of symptoms associated with chronic diseases such as Acquired Immunodeficiency Syndrome (AIDS), Alzheimer's disease, various types of depression and cancer (Wirleitner, Curr. Med. Chem. 2003, 10, 1581).

IDO activity also involves the development of age-related nuclear cataracts. IDO is the first enzyme in the biosynthesis of UV filters in the lens and is the rate-limiting enzyme. Ultraviolet filter compounds derived from tryptophan degradation (kynurenine and 3-hydroxykynurenine glucoside) modify the proteins present in the human lens. The amount of these ultraviolet filter compounds increases with age (Takikawa et al. Adv. Exp. Med. Biol. 1999, 241, 467) and it has been reported that these ultraviolet filter compounds cause the lens to become cloudy, which leads to the so-called Age-related nuclear cataract. IDO inhibitors block this natural process (Takikawa et al. Exp. Eye Res. 2001, 72, 271). IDO expression also involves inhibition of the immune response by preventing local T-lymphocyte proliferation. T-lymphocytes are very sensitive to the lack of tryptophan and in the absence of tryptophan, T-lymphocytes are arrested in the G1 phase of the cell cycle. This T cell-mediated inhibition of immune response is a factor that causes many diseases, including autoimmune diseases, allogeneic rejection, neurodegenerative disorders, depression, bacterial or viral infections (eg, human immunodeficiency virus HIV). And cancer (Swanson et al. Am. J. Respir. Cell Mol. Biol. 2003 30, 311). IDO inhibitors can be used to modulate T cell mediated immune responses.

 Most human tumors have been found to constitutively express IDO. Mouse tumor cells from pre-immunized mice have been shown to protect against rejection by expression of IDO, which is abolished by 1-MT administration. The efficacy of cancer treatment is then improved by the concomitant administration of IDO inhibitors (Uyttenhove et al. Nat. Med. 2003, 9, 1269). Pathological features of the tryptophan metabolism pathway, including infections of viruses such as AIDS, bacterial infections such as Lyme disease and streptococcal infection, neurodegenerative disorders (eg Alzheimer's disease, Huntington's disease and Paar) Jinsen disease), depression, cancer (including T-cell leukemia and colon cancer), eye diseases (such as cataracts and age-related yellowing), and autoimmune diseases. A variety of in vitro assays (Takikawa, et al. J. Biol. Chem. 1998, 263, 2041) can be used to screen (eg, high throughput screening), test reaction reference or IDO inhibitors of extracts obtained from natural sources. Activity, or determine its IDO inhibition kinetics constant.

 IDO is closely related to a variety of disease pathogenesis. It has been proven to be a target for major diseases such as cancer, Alzheimer's disease, depression, cataract, etc. IDO inhibitors have broad application prospects as drugs, but so far there is no suitable IDO inhibitors can be marketed as drugs, so it is of great theoretical and practical value to find new and effective IDO inhibitors. Existing studies have shown that the IDO inhibitor 1-MT (1-mercaptotryptophan) can enhance the sensitivity of tumor cells to immune stimulation of T cells in vitro; it can delay the growth of tumor cells and enhance chemotherapy drugs in animal models. Its anti-tumor effect, and it works for almost all spontaneous tumors, which brings new hope for tumor immunotherapy. 1-MT was included in the RAID (rapid access to intervention development) program by the National Cancer Institute, and entered the Phase I clinical trial in the fall of 2007. However, it is regrettable that most of the existing IDO inhibitors have low inhibitory efficacy. 1-MT is a commonly used IDO inhibitor in various in vitro and in vivo experiments, and its inhibition constant Ki is only 34 μΜ. Therefore, it has been found that new and highly effective IDO inhibitors have significant application value.

The tryptamine is an quinazoline alkaloid whose chemical name is 吲哚[ 2,1-b ]quinazoline-6,12-dione. Tryptophan is a yellow needle crystal which is mainly found in blue plants such as horse blue, indigo, and indigo (Honda G, et al. Planta Medica, 1980, 38(3): 275-276.). Alternatively, it can be extracted from the fermentation broth of the cockroach (Hosoe T, et al. Mycopathologia, 1999, 144(1): 9-12. ). In recent years, domestic and foreign scholars have carried out partial research on the pharmacology of tryptophan, and its pharmacological effects are mainly manifested in antibacterial, anti-inflammatory, anti-tumor and anti-parasitic (Earthur C, et al. Fitoterapia, 2005, 76(3- 4): 324-332; Motoki T, et al. Biol Pharm Bull, 2005, 28(2): 260-266). Although tryptamine can be extracted from the metabolites of blue plants and microorganisms such as indigo, malan, and indigo, the separation process is long and the extraction rate is low, which is difficult to meet the needs of research and clinical use. Only by exploring the artificial synthesis route which is short in time, high in yield and easy to obtain, can provide more resources for the application of tryptamine, making it possible for further development and application.

In recent years, pharmaceutical chemists have been working on the synthesis of tryptamine and its derivatives. The main method of synthesizing tryptamine is the reaction of hydrazine with isatoic anhydride. This method has a high yield and mild reaction conditions. In addition, functionalized groups can be introduced on both the ruthenium and isatoic anhydride precursors to synthesize various functional tryptamines. At present, the most important method for synthesizing hydrazine is to use hydration of trichloroacetaldehyde, hydroxylamine and aniline in an aqueous solution of hydrochloric acid to form a hydrazine compound, and then ring closure to obtain hydrazine under concentrated sulfuric acid. This method is very suitable for the synthesis of halogens. And the alkyl tryptone, the yield is also higher. However, this method is difficult to synthesize a tryptophan containing a reactive group because these groups are prone to various side reactions during the synthesis of hydrazine. There are two main methods for synthesizing isatoic anhydride. Method 1: Anthranilic acid reacts with triphosgene, and the method has high yield. However, the introduction of reactive groups on the starting materials is still very complicated. For example, using anthranilic acid as a raw material and introducing another amino group in the amino position, undergo a series of steps such as amino protection, nitro introduction, amino deprotection, Raney nickel reduction of nitro, etc., in order to obtain 2,5. - Diaminobenzoic acid. Method 2: Oxidation of ruthenium, that is, the use of H ₂ O ₂ -acetic anhydride system or Cr0 ₃ in acetic acid-acetic anhydride co-solvent to obtain isatoic anhydride, but this requires the synthesis of ruthenium. Therefore, the synthesis of functionalized tryptamine is a challenge to organic synthesis. Summary of the invention

 In order to overcome the deficiencies of the prior art, the present invention has been structurally modified to improve the solubility and pharmacological activity of tryptamine, with the aim of obtaining an active compound having application value. The research and pharmacological tests of the present invention show that the tryptophan derivative formed by introducing a triazole group into the tryptamine molecule can be used as a more efficient IDO inhibitor, which has antibacterial, anti-inflammatory, anti-tumor and the like. Pharmacological activity has broad application prospects. Further, the synthesis method of the present invention has the advantages of an operation unit, a mild condition, a high yield, and the like, and is easier to industrially produce than the conventional method for synthesizing a tryptophan derivative.

It is an object of the present invention to provide a tryptophan IDO having a 1,2,3-triazole structure. Inhibitors and methods for their preparation and use.

 The present invention relates to the compound, its tautomeric form, its structural analog or a pharmaceutically acceptable salt thereof, and a composition containing at least one of the compound, its structural analog or a pharmaceutically acceptable salt thereof. For use in inhibiting IDO, as well as in treating and/or preventing diseases having pathological features of the IDO-mediated tryptophan metabolism pathway. Such diseases include, but are not limited to, tumors, cancer, eye diseases, autoimmune diseases, mental disorders, depression, and anxiety disorders. Such uses include in vivo and in vitro applications, as well as in the preparation of pharmaceuticals, IDO inhibitors, and pharmaceutical compositions.

 The object of the present invention is achieved by the following technical solutions.

 In one aspect, the invention provides a compound of formula I or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or exosome thereof

Wherein R ¹ or R ^{2 is} independently selected from the group consisting of hydrogen, CC ₅ alkyl, a substituent of the formula II, and one of R ¹ and R ² is a substituent of the formula II,

Wherein R ^{3 is} selected from aryl; n = 0, 1, 2 or 3.

For the above compounds or pharmaceutically acceptable salts, solvates, polymorphs, enantiomers or racemic mixtures thereof, preferably, R ³ is phenyl, n = 1; more preferably, R ¹ and The other of R ² is fluorine or hydrogen; further preferably, R ¹ is fluorine or hydrogen, R ² is a substituent of the formula II, or R ¹ is a substituent of the formula II, and R ² is fluorine or hydrogen.

 The structure of the most preferred compounds of the invention is as follows:

The invention also provides a process for the preparation of the above compound or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof, the process comprising the steps of: in the presence of a base, formula XIII a compound of the formula XIV I,

 XIII XIV.

 Preferably, the above preparation method comprises the following steps:

(1) A compound of formula III is reacted with NaN ₃ in disulfoxide to form a compound of formula IV.

 III IV

(2) reacting a compound of formula IV with a compound of formula V in the presence of sodium ascorbate and cuprous iodide to form

 V VI

(3) a compound of formula VI is reacted with a reduced iron powder to form a compound of formula VII

 VII

 (4) The compound of formula VII is reacted with hydrated trichloroacetaldehyde and hydroxylamine hydrochloride in the presence of anhydrous sodium sulfate to form formula VIII.

 VIII

(5) reacting a compound of the formula VIII in concentrated sulfuric acid in the presence of anhydrous sodium sulfate to form a compound of the formula IX;

 IX

 (6) reacting a compound of the formula IX with a compound of the formula X in the presence of triethylamine to form a compound of the formula I,

 X

Wherein, in Formula I, R ^{1 is} selected from the group consisting of hydrogen, CC ₅ alkyl, and halogen, and R ² is a substituent of Formula II;

 (7) In the presence of chromium trioxide, the compound of formula VIII is reacted in glacial acetic acid and acetic anhydride to form a compound of formula XI.

(8) reacting a compound of the formula XI with a compound of the formula XII in the presence of triethylamine to form a compound of the formula I,

 XII

Wherein, in Formula I, R ¹ is a substituent of the formula II, and R ^{2 is} selected from the group consisting of hydrogen, C _r C ₅ alkyl and halogen.

 More preferably, the above preparation method comprises the following steps:

(1) The compound of the formula III is reacted with NaN ₃ at room temperature for 0.5-1 hour at room temperature. After completion of the reaction, water is added, and the mixture is extracted with ethyl acetate. The ethyl acetate phase is washed with water and dried over anhydrous sodium sulfate. An ester, that is, a compound of the formula IV;

 (2) The compound of the formula IV is reacted with sodium ascorbate, cuprous iodide, acetonitrile, water and the compound of the formula V at room temperature under a nitrogen atmosphere. After the reaction is completed, the reaction product is poured into water, and acetic acid B is used. The ester is extracted, and the saturated brine is washed until neutral. The ethyl acetate phase is dried over anhydrous sodium sulfate to remove ethyl acetate.

 (3) The compound of the formula VI is reacted in a 5% by weight aqueous solution of ammonium chloride at 80 ° C for 4-5 hours in the presence of activated reduced iron powder. After the reaction, it is cooled to room temperature and adjusted by sodium carbonate. The pH was 8-9, and the mixture was stirred with ethyl acetate for 0.5 hr, then filtered over Celite. That is, a compound of the formula VII;

 (4) The compound of the formula VII is stirred and mixed with hydrated trichloroacetaldehyde, anhydrous sodium sulfate and water, and then a 5% by weight aqueous solution of hydrochloric acid, hydroxylamine hydrochloride and water are successively added, and the reaction is carried out at 100 ° C for 2-3 hours. After the reaction is completed, it is cooled to room temperature, suction filtered, and the filter cake is washed with 10% by weight aqueous hydrochloric acid and water, and dried under vacuum to obtain a compound of formula VIII;

 (5) Dissolving the compound of the formula VIII in concentrated sulfuric acid at normal temperature and vigorous stirring, and then raising the temperature to 65 ° C for 4-5 hours. After the reaction is completed, the reaction solution is poured into cold water to precipitate a solid, and suction filtration is carried out. The filter cake is washed with water and dried under vacuum to obtain a compound of the formula IX;

 (6) The compound of the formula IX is reacted with the compound of the formula XII in terpene at 110 ° C for 3-4 hours in the presence of triethylamine. After completion of the reaction, the triethylamine and toluene are removed, and anhydrous ethanol is added. Recrystallization, that is, the compound of formula I,

Wherein, in Formula I, R ^{1 is} selected from the group consisting of hydrogen, C _r C ₅ alkyl, and halogen, and R ² is a substituent of Formula II;

(7) The compound of formula VIII is in glacial acetic acid and acetic anhydride in the presence of chromium trioxide After reacting at 80-90 ° C for 3 hours, after the reaction is completed, the reaction product is cooled to room temperature, water is added, suction filtration, and the solid is washed with water, and filtered to obtain a compound of the formula XI;

 (8) The compound of the formula XI is reacted with the compound of the formula XII in terpene at 110 ° C for 3 hours in the presence of triethylamine. After the reaction, the triethylamine and toluene are removed, and then ethanol is recrystallized. That is, the compound of formula I,

Wherein, in Formula I, R ¹ is a substituent of the formula II, and R ^{2 is} selected from the group consisting of hydrogen, C _r C ₅ alkyl and halogen.

For the above preparation method, preferably, in the step (1), the molar ratio of the compound of the formula III to NaN ₃ is 1:1.2; preferably, in the step (2), The molar ratio of the compound of the formula IV to the sodium ascorbate, cuprous iodide and phenylacetylene is 1:

0.4: 0.2: 2; Preferably, in the step (3), the molar ratio between the compound represented by the formula VI and the reduced iron powder is 1:8; preferably, in the step (4) The molar ratio of the compound of the formula VII to the hydrated trichloroacetaldehyde, anhydrous sodium sulfate, and hydroxylamine hydrochloride is 1: 1: 1: 3; preferably, in the step (6) The molar ratio of the compound of the formula IX to the compound of the formula XII and the triethylamine is 1: 1:5; preferably, in the step (7), the chromium trioxide The molar ratio between the compound represented by the formula VIII is 1: 1.1-1.2, and the molar ratio between the glacial acetic acid and acetic anhydride is 1:1; preferably, in the step (7), The molar ratio of the compound of the formula XI to the compound of the formula XII and triethylamine is 1: 1: 4-6.

 In another aspect, the invention provides the use of a compound of the above, or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof, in the manufacture of an IDO inhibitor.

 Further, the present invention provides a compound or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof for the preparation of a prophylactic and/or therapeutic treatment with indole-2,3-bis Oxygenase-mediated application of drugs for diseases associated with tryptophan metabolism disorders. Preferably, the disease associated with a guanamine-2,3-dioxygenase-mediated disorder of tryptophan metabolism is selected from the group consisting of a tumor, a cancer, an Alzheimer's disease, an autoimmune disease, a cataract, a mental disorder. , one or more of depression and anxiety.

 In still another aspect, the present invention provides a pharmaceutical composition for a guanamine-2,3-dioxygenase inhibitor, which comprises the above compound or a pharmaceutically acceptable salt, solvate thereof, Polymorphs, enantiomers or racemic mixtures, and pharmaceutically acceptable carriers and/or excipients.

In still another aspect, the present invention provides a method for treating, preventing or delaying a disease associated with a guanamine-2,3-dioxygenase-mediated disorder of tryptophan metabolism, the method comprising administering A therapeutically effective amount of the above compound or a pharmaceutically acceptable salt thereof, solvate in a patient in need of treatment , a polymorph, an enantiomer or a racemic mixture or a pharmaceutical composition as described above,

 Preferably, the disease associated with a guanamine-2,3-dioxygenase-mediated disorder of tryptophan metabolism is selected from the group consisting of a tumor, a cancer, an Alzheimer's disease, an autoimmune disease, a cataract, a mental disorder. , one or more of depression and anxiety.

In one embodiment of the present invention, the present invention provides an 8-amino-1,2,3-triazole-containing tryptamine IDO which inhibits the structure of the IDO inhibitor as follows:

 Wherein R is hydrogen or halogen.

 In a specific embodiment, the 8-position tryptophan IDO inhibitor having a 1,2,3-triazole structure

 The synthetic route of the above 8-position tryptamine IDO inhibitor containing 1,2,3-triazole structure is as follows:

 Specifically, the preparation method includes the following steps:

(1) Synthesis of 1-azidoindol-4-nitrobenzene The p-nitrobenzyl chloride, sodium azide and disulfoxide were sequentially added to the reaction flask, and the aluminum foil was protected from light at room temperature for 0.5-1 hour. The reaction was confirmed by thin layer chromatography (TLC). Water is added to the system, and the mixture is extracted with ethyl acetate. The organic phase is washed with water, dried with anhydrous sodium sulphate, and the ethyl acetate is removed by rotary evaporation under vacuum. The molar ratio of p-nitrobenzyl chloride to sodium azide is 1: 1.2;

 (2) Synthesis of 1-(4-nitrobenzyl)-4-phenyl-1H-1,2,3-triazole

 The 1-azidoindolyl-4-nitrobenzene, sodium ascorbate, cuprous iodide, acetonitrile, water and phenylacetylene prepared in the step (1) are sequentially added to the reaction flask, protected by nitrogen, and stirred at room temperature. After overnight, the TLC test showed that the reaction was completed, the reaction mixture was poured into water, extracted with ethyl acetate, and washed with saturated brine, and the organic phase was dried over anhydrous sodium sulfate. That is, wherein, the molar ratio of 1-azidoindol-4-nitrobenzene, sodium ascorbate, cuprous iodide to phenylacetylene is 1: 04: 0.2: 2;

 (3) Synthesis of 4-(4-phenyl-1H- 1,2,3-triazol-1-yl)nonylaniline

 Adding reduced iron powder, 5% by weight aqueous solution of ammonium chloride to the reaction flask, heating the reaction solution to 100 ° C, activating the iron powder for 1 hour, then cooling to 80 ° C, slowly adding the batch step (2) Preparation of 1-(4-nitrobenzyl)-4-phenyl-1H-1,2,3-triazole, reaction 4-5 hours, TLC detection showed that after the reaction was completed, cooled to room temperature, added solid carbonic acid The pH of the sodium was adjusted to 8-9, and the mixture was stirred for 20 hours with 20 ml of ethyl acetate. The mixture was filtered over Celite, and the filtrate was extracted with ethyl acetate. The mixture was washed with saturated brine and dried over anhydrous sodium sulfate. Ethyl acetate, that is, wherein; the molar ratio of 1-(4-nitrobenzyl)-4-phenyl-1H-1,2,3-triazole to reduced iron powder is 1:8;

 (4) Preparation of an anthracene derivative containing 1,2,3-triazole

 To the reaction flask, hydrated trichloroacetaldehyde, anhydrous sodium sulfate and water were added, and 4-(4-phenyl-1H-1,2,3-triazole-1-) prepared by the step (3) was added under stirring. Base mercaptoaniline, then adding 5% by weight aqueous hydrochloric acid solution, then adding hydroxylamine hydrochloride, water, and then raising the temperature to 100 ° C for 2-3 hours. After TLC detection showed that the reaction was completed, the reaction solution was cooled to room temperature. After suction filtration, the filter cake is washed with 10% by weight aqueous hydrochloric acid solution, washed with water and dried under vacuum to obtain 4-(4-phenyl-1H-1,2,3-triazol-1-yl group. The molar ratio of mercaptoaniline to hydrated trichloroacetaldehyde, anhydrous sodium sulfate, hydroxylamine hydrochloride is 1: 1: 1: 3;

 (5) Preparation of an isatin derivative containing 1,2,3-triazole structure

 The concentrated sulfuric acid is added to the reaction flask, and the product obtained in the step (4) is added in portions under vigorous stirring, until all is dissolved in concentrated sulfuric acid, and then the temperature is raised to 65 ° C, and the reaction is carried out for 4-5 hours. The TLC detection shows the reaction of the raw materials. After completion, the reaction solution is poured into cold water, that is, a yellow solid is precipitated, which is suction filtered, and the filter cake is washed with water and dried in vacuo;

(6) Synthesis of 8-aminotryptamine IDO inhibitors containing 1,2,3-triazole structure The product obtained in the step (5) and the isatoic anhydride derivative represented by the following formula are added to the reaction flask, then triethylamine and terpene are added, the temperature is raised to 110 ° C, and the reaction is stirred under reflux for 3-4 hours, and TLC is detected. After the reaction is completed, the triethylamine and toluene are removed by vacuum rotary evaporation, and recrystallized by adding anhydrous ethanol to obtain the isoflavone derivative having the 1,2,3-triazole structure prepared in the step (5). The molar ratio of the substance to the isatoic anhydride derivative and triethylamine is 1: 1:5.

 Wherein R is hydrogen or halogen.

In another embodiment of the present invention, the present invention provides a 2-position tryptamine-containing IDO having a 1,2 azole structure as follows:

 Wherein R is hydrogen or halogen.

In a specific embodiment, the above formula for the tryptophan IDO inhibitor having a 1,2,3-triazole structure at the 2-position is as follows:

 The synthetic route of the above-mentioned 2-position 1,2,3-triazole-containing tryptophan IDO inhibitor is as follows:

Specifically, the preparation method includes the following steps:

 (1) Synthesis of isatoic anhydride containing triazole structure

Adding glacial acetic acid and acetic anhydride to the reaction flask, and then adding the above-obtained isoflavone derivative containing 1,2,3-triazole structure in portions, heating the reaction solution to 80-90 ° C, and then batching Adding trioxane Chromium, continue to react for 3 hours, TLC test shows that the reaction of the starting material is complete, the reaction solution is cooled to room temperature, 1.5 mL of water is added, suction filtration, the solid is washed with a large amount of water, and the oil pump is pumped to obtain a green solid without separation, leaving the next step reaction;

 (2) Synthesis of a tryptophan IDO inhibitor containing 2-, 2,3-triazole structure

The compound represented by the following formula, the isocyanuric acid anhydride-containing phthalic anhydride, the terpene benzene, and the triethylamine prepared in the step (1) are sequentially added to the reaction flask, and then the temperature is raised to 110 ° C for 3 hours, and the reaction is completed. After that, the triethylamine and the terpene are first removed, and then recrystallized by adding ethanol to obtain a dark green solid.

 Wherein R is hydrogen or halogen.

 IDO has been shown to be closely related to a variety of human major diseases such as Alzheimer's disease, cataract, cancer, etc. Experiments have shown that the 1,2,3-triazole-containing tryptophan derivatives of the present invention have been Known IDO inhibitors have more potent IDO inhibition and can be used to treat diseases with IDO-mediated pathological features of the tryptophan metabolism pathway, and have antibacterial, anti-inflammatory, anti-tumor, autoimmune and other Pharmacological activity, therefore, the novel IDO inhibitor obtained by the invention can be used as a novel drug and has broad application prospects. At the same time, it can be used as a further modified pharmaceutical intermediate with potential application value for the development of other new drugs.

 The method for preparing the above-mentioned tryptophan IDO inhibitor containing 1,2,3-triazole structure has the advantages of convenient operation, mild reaction conditions, solvent saving, pollution reduction and the like, and is convenient for industrial production. Specifically, the preparation method of the present invention uses p-nitrobenzyl chloride as a raw material, reacts with sodium azide to form 1-azidoindol-4-nitrobenzene, and then 1,3-dipolar cycloaddition with phenylacetylene. The reaction is formed to form a triazole compound. The compound is reacted with an amino group in an ammonium chloride solution in the presence of a reduced iron powder; then, the amino compound reacts with hydrated trichloroacetaldehyde or hydroxylamine hydrochloride to form a triazole-containing hydrazine. Then, in the presence of concentrated acid, the ruthenium ring is closed to form a triazole-containing eosin derivative. The isoxazole derivative-containing isatin derivative is reacted with isatoic anhydride or a derivative thereof under alkaline conditions in a solvent of toluene to synthesize a series of tryptamine derivatives containing a triazole structure. The best way to implement the invention

 The present invention is further described in detail with reference to the preferred embodiments thereof.

The experimental methods in the following examples are conventional methods unless otherwise specified. Following implementation The raw materials, reagent materials, etc. used in the examples can be purchased from conventional biochemical reagent stores or pharmaceutical companies without special instructions. Example 1: 8-(4-phenyl-lH-1,2,3-triazol-1-ylindenyl)tryptamine

(1) Preparation of 1-azidoindol-4-nitrobenzene

 p-Nitrobenzyl chloride (162 mg, 1 mmol), sodium azide (120 μ, 1.2 mmol), and diterpene (121 mg, 1.2 mmol) were sequentially added to the reaction flask. The aluminum foil was protected from light and reacted at room temperature for 0.5- Lh, TLC detection shows that after the reaction is completed, water is added to the reaction system, and the mixture is extracted with ethyl acetate. The organic phase is washed with water, dried over anhydrous sodium sulfate, and evaporated in vacuo to remove ethyl acetate to obtain a pale yellow-brown liquid. Next reaction;

(2) Preparation of 1-(4-nitrobenzyl)-4-phenyl-lH-1,2,3-triazole

 1-azidodecyl-4-nitrobenzene (178 mg, lmmol), sodium ascorbate (79 mg, 0.4 mmol), 4 匕4 (38 mg, 0.2 mmol), B (3 mL), water (0.3mL) and phenylacetylene (204mg, 2mmol) were added to the reaction flask in turn, protected by nitrogen, and stirred at room temperature overnight. After TLC detection showed that the reaction was completed, the reaction solution was poured into water, extracted with ethyl acetate, and washed with saturated brine. Neutral, the organic phase was dried over anhydrous sodium sulfate, and ethyl acetate was evaporated in vacuo.

The characterization data is as follows: ^-NMR (400 MHz, CDC1 ₃ ): δ = 8.24 (d, 2 Η), 7.83 (d, 2H) _: 7.75 (s, 1H), 7.44 (m, 4H), 7.35 (m, 1H) ), 5.71(s, 2H).

(3) Synthesis of 4-(4-phenyl-lH-1,2,3-triazol-1-yl)nonylaniline

Reduced iron powder (448mg, 8mmol), ammonium chloride (350mg, 6.54mmol), water (7mL) were added to the reaction flask, the reaction solution was heated to 100 °C, the iron powder was activated for 1h, and then cooled to 80 °C. The product of the above step (280 mg, 1 mmol) was slowly added in portions, and the reaction was carried out for 4-5 h. After the TLC test showed that the reaction was completed, it was cooled to room temperature, adjusted to pH = 8-9 by adding solid sodium carbonate, and stirred with 20 ml of ethyl acetate for 0.5 h. The celite was filtered, and the filtrate was extracted with ethyl acetate. The mixture was washed with saturated brine and dried over anhydrous sodium sulfate. The color solid was 228 mg, yield 91%.

The characterization data is as follows: ^-NMR (400 MHz, CDC1 ₃ ): δ = 7.79 (d, 2 Η), 7.60 (s, IH) 7.39 (t, 2H), 7.30 (t, IH), 7.13 (d, 2H) , 6.68(d, 2H), 5.44(s, 2H).

Azoazole structure

 To the reaction flask was added hydrated trichloroacetaldehyde (165 mg, 1 mmol), sodium sulphate (141 mg, 1 mmol), water (2.2 mL), and the above product (250 mg, 1 mmol) was added with stirring, then added (0.7 mL) 5% hydrochloric acid solution, then add hydroxylamine hydrochloride (209mg, 3mmol), water (0.95mL), then warm to 100 ° C, reaction 2-3h, TLC detection shows that after the reaction is completed, the reaction solution is cooled to room temperature, pumping Filtration, the filter cake was washed with 10% aqueous hydrochloric acid, washed with water and dried in vacuo to give 161 mg of pale yellow solid.

(5) Preparation of an isatin derivative containing a 1,2,3-triazole structure

 Concentrated sulfuric acid (2.5 mL) was added to the reaction flask, and the above step product (321 mg, 1 mmol) was added portionwise under vigorous stirring, until all was dissolved in concentrated sulphuric acid, and then the temperature was raised to 65 ° C, and the reaction was carried out for 4-5 h. After the TLC test showed that the reaction of the starting material was completed, the reaction mixture was poured into cold water to precipitate a yellow solid, which was filtered with suction. The filter cake was washed with water and dried in vacuo to give 289 mg of an orange solid, yield 95%.

 The characterization data is as follows: ifi-NMR (400 MHz, CDC13): δ = 7.97 (s, IH), 7.80 (d, 3H), 7.73 (s, IH), 7.61 (s, IH), 7.55 (d, IH) , 7.50(d, 2H), 6.93(d, IH), 5.55(s, 2H).

(6) Synthesis of 8-aminotryptamine IDO inhibitors containing 1,2,3-triazole structure

 The isoflavone derivative (304 mg, lmmol) containing the 1,2,3-triazole structure prepared in the step (5) and the isatoic anhydride (purchased from Aladdin Reagent Co., Ltd.) (163 mg, lmmol) were added to the reaction. In the bottle, triethylamine and terpene were added, and the temperature was raised to 110 ° C. The reaction was stirred under reflux for 3-4 h. After TLC detection showed that the reaction was completed, triethylamine and toluene were removed by vacuum rotary evaporation, and recrystallized from anhydrous ethanol. Finally, a yellow-green solid 284 mg was obtained with a yield of 70%.

The characterization data is as follows: ifi-NMR (400 MHz, CDC1 ₃ ): δ = 8.66 (d, IH), 8.44 (d, IH), 8.03 (d, IH), 7.80 (m, 4H), 7.76 (m, 2H) ), 7.69(t, IH), 7.43(t, 2H), 7.34 (t, 1H), 5.68(s, 2H). Example 2: 2-Fluoro-8-(4-phenyl-lH-1,2,3-triazol-1-ylindenyl)tryptamine

 The isoflavone derivative containing the 1,2,3-triazole structure prepared in the step (5) of Example 1 (304 mg, lmmol) and 5-fluoroindigonic anhydride (purchased from Yancheng City Medico chemical production limited ( 181mg, lmmol ) was added to the reaction flask, then added triethylamine, toluene, and the temperature was raised to 110 ° C, and the reaction was stirred for 4 hours under reflux. After TLC detection showed that the reaction was completed, triethylamine and toluene were removed by vacuum rotary evaporation. It was recrystallized by adding absolute ethanol, and finally a dark yellow-green solid 275 mg was obtained in a yield of 65%.

The characterization data is as follows: ifi-NMR (400 MHz, CDC1 ₃ ): δ = 8.65 (d, 1 Η), 8.07 (m, 2H), 7.82 (m, 5H), 7.57 (m, 1H), 7.43 (t, 2H) ), 7.35(t, 1H), 5.69(s, 2H). Example 3: Preparation of 8-fluoro-2-(4-phenyl-1H-1,2,3-triazolylhydrazyl)tryptamine

Synthesis of 1 triazole structure of isatoic anhydride

 0.42 mL of glacial acetic acid and 0.42 mL of acetic anhydride were added to a 25 mL reaction flask, and then (152 mg, 0.5 mmol) of a triazole-containing ruthenium (see Example 1) was added in portions, and the reaction solution was heated to 80- At 90 °C, the chromium trioxide was added in batches and the reaction was continued for 3 hours. After the TLC test showed that the reaction of the raw materials was complete, the reaction solution was cooled to room temperature, 1.5 mL of water was added, suction filtration was applied, and the solid was washed with a large amount of water, and the oil pump was drained. The green solid does not have to be separated and is left to the next reaction.

-tryptamine I containing a -triazole structure

 Add 5-fluoroindole in order to the reaction bottle (purchased from Wuhan Shisheng Technology Development Co., Ltd.)

(165 mg, 1 mmol), the triazole-containing isatoic anhydride (320 mg, 1 mmol) prepared in the step (1), 2.5 ml of toluene, triethylamine (505 mg, 5 mmol), and then heated to 110 ° C for 3 h. After the end of the reaction, triethylamine and terpene were removed, and then ethanol was recrystallized to give a dark green solid. The yield was 65%. The characterization data is as follows: ^-NMR (400 MHz, CDC1 ₃ ): δ = 8.65 (m, 1H), 8.42 (s, 1H) _: 8.06 (d, 1H), 7.80 (m, 4H) 7.61 (m, 1H) , 7.52 (m, 1H), 7.44 (t, 2H), 7.36 (t, 1H), 5.80 (s, 2H). Example 4: Detection of IDO inhibitory activity

 The construction of the plasmid containing the human IDO gene, expression, extraction and purification in E. coli were carried out according to the method of Littlejohn et al. (Takikawa 0, Kuroiwa T, Yamazaki F, et al. J. Biol. Chem. 1988, 263, 2041-2048). The inhibitory activities of the isolated components and monomeric compounds against IDO were examined by the methods described below.

 50 mM potassium citrate buffer (pH 6.5), 40 mM vitamin C, in a 96-well plate.

400 g/ml catalase, 20 μM of hydrazinyl blue and IDO enzyme were mixed. The substrate L-tryptophan and the sample to be tested were added to the above mixture. The reaction was carried out at 37 ° C for 60 minutes, and the reaction was terminated by the addition of 30% (w/v) trichloroacetic acid. The 96-well plate was heated at 65 °C for 15 minutes to complete the conversion from decanoyl uric acid to kynurenine, followed by rotation at 6000 rpm for 5 minutes. Remove the ΙΟΟμΙ supernatant from each well and transfer to a new 96-well plate. Add 2% (w/v) p-dimercaptoaminobenzaldehyde in acetic acid. The yellow color produced by the reaction of kynurenine can be used. The microplate reader was observed at 490 nm. The preliminary test showed that the tryptone derivatives prepared in Examples 1-3 had IDO inhibitory activity. Example 5: Whether it is a reversible inhibitor

 At a fixed inhibitor concentration, a series of different concentrations of enzyme are reacted with the inhibitor and the rate of reaction is determined. The reaction rate was plotted against the enzyme concentration (V ~ [E]), and it was determined according to the characteristics of the curve whether it is a reversible inhibitor.

 Reaction conditions: In a 500 μΐ reaction system, first add 50 mM potassium phosphate buffer (pH 6.5), 40 mM vitamin C, 400 g/ml catalase, 20 μM ruthenium blue, 300 mM substrate L-color ammonia. Acid or simultaneous addition of 100 mM inhibitor, the mixture was incubated at 37 ° C for 5 minutes, and then added different volumes of IDO enzyme to the mixture, the reaction was carried out at 37 ° C for 30 minutes, added 30% (w / v) three The reaction was terminated by chloroacetic acid 200 μl, and the reaction system was heated at 65 ° C for 15 minutes to complete the conversion from decanoyl kynurenine to kynurenine, then rotated at 12000 rpm for 10 minutes, and the supernatant was taken up to an equal volume of 2%. (w/v) Mix the acetic acid solution of dimercaptoaminofurfural and measure the reading at 490 nm with a microplate reader. The test results showed that the tryptamine derivatives prepared in Examples 1-3 were reversible IDO inhibitors. Example 6: Determination of inhibitor type and determination of Ki value

In a 500 μl reaction system, first add 50 mM potassium phosphate buffer (pH 6.5), 40 mM vitamin C, 400 g/ml catalase, 20 μM yttrium blue, and add 100, 250, respectively. 300 mM substrate L-tryptophan, at different substrate concentrations, different concentrations of compounds were added to each tube reaction system, the mixture was incubated at 37 ° C for 5 minutes, and then ΙΟμΙ IDO was added to the mixture. 20nM), the reaction was carried out at 37 ° C for 30 minutes, 30% (w / v) trichloroacetic acid 200μ1 was added to terminate the reaction, and the reaction system was heated in a water bath at 65 ° C for 15 minutes to complete the chymolyserine To the conversion of kynurenine, then centrifuge at 12000 rpm for 10 minutes, take the supernatant and mix with an equal volume of 2% (w/v) acetic acid solution of diammonium aminobenzaldehyde, and measure the wavelength of 490nm with a microplate reader. reading. The type of inhibitor of the compound was determined by Dixon's mapping method (l/v~[I]), and the Ki value of the inhibitor was obtained by plotting S/v~[I]. Example 7: Half effective inhibition concentration IC ₅ . Determination of (in vitro)

50 mM potassium phosphate buffer (pH 6.5), 40 mM vitamin C, 400 g/ml catalase, 20 μM sulfhydryl blue, substrate L-tryptophan 150 mM and an inhibitor were first mixed. The concentration of the inhibitor was selected from 100, 200, 400, 600, 800, 1000, and 1200 μM, and the mixture was incubated at 37 ° C for 5 minutes, and then the IDO enzyme was added to the above mixture. The reaction was carried out at 37 ° C for 30 minutes, 30% (w / v) trichloroacetic acid 200 μl was added to terminate the reaction, and the reaction system was heated at 65 ° C for 15 minutes to complete the reaction from decanoyl kynurenine to guanine ammonia. Acid conversion, then rotate at 12000 rpm for 10 minutes, take 200μ1 supernatant and mix with an equal volume of 2% (w/v) of di-aminobenzaldehyde in acetic acid solution. The yellow color produced by the reaction of kynurenine can be used. The microplate reader was tested at 490 nm and the results were obtained using IC ₅ . Calculation software calculation (see Table 1 for results). The measurement mechanism is as follows:

30% TCA, 65 ° C, 15 min

λ max = 490 nm Example 8: Half effective inhibition concentration IC ₅ . Determination of (cell)

Plasmid pcDNA3.1-MDO was transiently transfected into HEK 293 cells using liposome Lipofectamin 2000. At the time of cell level inhibitor activity assay, HEK293 cell culture medium was high glucose DMEM containing 50 U/mL penicillin, 50 U/mL streptomycin, 10% FBS, 37 °C, 5% CO ₂ culture. After the cells were transfected into the plasmid for 24 h, the drug to be tested was added. After incubation for a period of time, the supernatant was taken. In another 96-well plate, ΙΟμ 30% (w/v) trichloroacetic acid was added and heated at 65 °C for 15 min to complete the conversion of decanoyl kynurenine to kynurenine, and then centrifuged at 12000 rpm for 10 min. Take an equal volume of 2%. (w/v) mixed color development of diacetamidofuraldehyde in acetic acid solution, and finally the absorbance was measured at 490 nm using a microplate reader.

 Using the methods of Examples 5-8 above, the IDO inhibitory activity of the compounds prepared in Examples 1-3 was determined, and the IDO inhibitor 1-mercaptotryptophan (1-MT, which is commonly used in current and in vitro experiments) was used. As a control, the measurement results are shown in Table 1. Table 1 Examples Results of IDO inhibitory activity of compounds synthesized in 1-3

Example 9: Effect of anti-mouse Ρ388 leukemia

 Fine package

 Mouse lymphocytic leukemia cell line Ρ388, purchased from Nanjing Kaiji Biotechnology Co., Ltd.

 2. Experimental animals

 6-8 weeks old DBA/2 mice, inbred lines, SPF grade, 50, male and female, body mass (22.0 ± 1.6) g, purchased from Shanghai Slack Laboratory Animal Co., Ltd. All mice were housed in the Barrier Environmental Animal Laboratory of the Experimental Animal Center of Shenyang Pharmaceutical University.

 3. P388 leukemia mouse modeling method

P388 leukemia cells in logarithmic growth phase cultured in vitro were washed twice with physiological saline, and then intraperitoneally inoculated into 2 DBA/2 mice for about 1×10 ⁶ cells, and sterilized on the 8th day after sacrifice. Under the ascites, the milky white translucent, set the density of the cells in a sterile container with physiological saline to 5 x l0 ⁶ / mL, take a little suspension for trypan blue staining, count under light microscope, the number of viable cells should be more than 95%, then the above tumor cell suspension was intraperitoneally injected (ip) 0.2 mL (about 1 x 10 ⁶ cells) into each mouse under sterile conditions, and a total of 50 cells were inoculated.

 4. Risk grouping, drug administration and test method

Prepared in 20% DMSO, diluted to a suitable concentration of working solution according to the body weight of the mice according to the weight of the mice, the maximum concentration of DMSO is not more than 10%. 50 successfully vaccinated mice were randomly divided into 5 groups, namely blank control group, solvent control group and tryptamine derivative group (Examples 1, 2, 3, each group). 35mg/kg), 10 in each group, half male and half female. The administration started on the next day of inoculation. On the 1st to 7th day of the experiment, the blank control group, each mouse ip normal saline 0.2 mL / d; solvent control group, each mouse ip 10% DMSO 0.2 mL / d; tryptamine derivative group ( In the first, second, and third groups, each mouse ip corresponds to a drug of 35 mg/kg, and the final volume is 0.2 mL.

 Calculation method of life extension rate of leukemia mice: The survival period of tumor-bearing mice is calculated from the day of inoculation. Life extension rate = (average survival of the experimental group - average survival of the control group) / average survival time of the control group X 100%

 5. Results

 Table 2

 Number of groups (n) Average number of days of survival (d) Life extension rate (%) Blank control group 10 12.40 ± 1.43 - Solvent control group 10 13.30 ± 1.83 - Example 1 group 10 20.60 ± 1.78 54.88 Example 2 group 10 21.30 ± 1.09 60.15 Example 3 Group 10 19.50±1.84 46.62 The mice in the two control groups grew rapidly in ascites. On the 8th day after inoculation, the abdomen was able to see obvious bulging, and the movement was slow. After death, a large amount of turbid bloody ascites was seen in the laparotomy. It can be seen that more gray-white and fragile tumor masses are attached. After the administration of the mice in each of the drug-administered groups, the abdomen gradually swelled. In the three groups, abdominal augmentation was observed around 12 days. After the death, a large amount of bloody ascites was also observed in the abdominal cavity, and the tumor mass was less.

 6. Discussion

 According to the results, the tryptophan derivatives of the present invention can significantly prolong the survival of P388 leukemia mice, indicating that they have a good anti-tumor effect in P388 leukemia mice. Example 10: Effect on learning and memory ability of Alzheimer's rats

 Main reagents and instruments

 Quinolinic acid (QA): supplied by Sigma. Kit (Tunnel): Provided by Wuhan Boster Bioengineering Co., Ltd., product number MK1020. Brain stereo locator NARISHIGE SN-2 type. Morris Water Maze: Manufactured by the Department of Pharmacology, China Medical University.

 2. Experimental animals

 Fifty male Wistar rats, half male and half female, weighing (280 ± 10) g, were provided by the Experimental Animal Center of the Chinese Medical University.

 3. Animal grouping, model establishment and medication

 50 rats were randomly divided into 5 groups, namely, the sham injury group, the model group, and the tryptamine derivative group (Examples 1, 2, and 3, 50 mg/kg each), 10 rats in each group, and the weight of the rats between the groups. There was no statistical difference.

Except the blank group, the experimental animals were anesthetized with 3% pentobarbital sodium 1 ml/kg ip; fixed on the brain stereotaxic instrument, the cranial cranial midline incision, reference to the rat brain stereotactic map (AP 0.2 mm, ML 2.5mm, DV 4.5mm), after stereotactic positioning of the bilateral hippocampal CA1 area, drill the skull, use a 2μ1 micro-injector to vertically insert the needle, and dissolve QA 2μ1 (containing 150nmol QA) with 0.01mol/L (pH 7.4) PBS buffer. Slowly injected into the bilateral hippocampal CA1 area, the dummy injury group was injected with 0.01mol/L (pH7.4) PBS 2μ1, the injection time per side was 5min, the needle was kept for 5min, the skull was sealed with dental mud after needle extraction, and the skin was partially disinfected after disinfection. Intramuscular injection of penicillin for 3 days to prevent infection. One week after operation, each group was dosed. The sham injury group and the model group were given distilled water 2 ml/supply for 2 weeks.

 Morris water maze test method

 After 1 week of treatment, each rat's head and neck hair was blacked out with a common hair dye. The laboratory temperature was 24~25 °C during the test, and 1 kg of milk powder was added to the water maze, and it was washed with hot water. Add water to the safety platform lcm, and keep the water temperature at around 22 °C. Connect the installed camera unit to the computer monitor and printer. The pool is marked with four water inlets from east to west and north and south. The pool is divided into four quadrants, SW, NW, SE, NE. The safety platform is located in the SW quadrant, 23cm from the center of the circle. After the safety platform is fixed, it is separated from the four quadrants. The rats were placed in the water pool, and the time (latency, SPL) and number of inductions of the rats from the water inlet point were recorded. If the animal could not find a safety platform within 2 minutes, the experimenter put them back to safety. Platforms, movement paths and other indicators, as a positioning of academic achievement. On the first day, the rats were allowed to swim freely for 5 min to familiarize themselves with the environment. The next day, the first day, each training session was 4 times, for 3 consecutive days, and the fifth day was carried out for space exploration experiments (ie, removing the safety platform and observing the mouse within 2 minutes) The ratio of swimming distance to total distance in the quadrant) to detect spatial memory in rats. Each group of 5 groups was performed in parallel.

 4, the experimental results

 table 3

 Group latency ( s ) distance ( cm ) % of induction induction % sham injury group 41.98 ± 10.52 682.62 ± 323.35 4.50 ± 1.51 3.4 ± 0.4 model group 70.87 ± 9.62 977.13 ± 111.10 1.33 ± 0.89 15.1 ± 1.3 Example 1 group 41.39 ± 6.63* 690.87±108.69* 2.50±1.24* 9.1±0.3* Example 2 Group 37.16±10.18* 710.50±230.13* 2.75±1.42* 8.3±1.4* Example 3 Group 40.21±9.58* 705.42±198.34* 2.68±1.37* 8.6±0.8*

* Compared with the model group, P O.05 or 0.01

As can be seen from Table 3, the effects of tryptamine derivatives (Examples 1, 2, 3) on learning and memory processes induced by QA in hippocampus-induced AD rats (positioning navigation test): the treatment group has a lower latency than the model group, and the navigation path is shortened. The number of inductions decreased and the percentage of induction decreased (x 2 test, P < 0.05). Effects of tryptamine derivatives (Examples 1, 2, 3) on the memory intensity of hippocampus-induced AD rats with QA injury (spatial exploration test): The quadrants of the swimming routes of the model group were almost average, still blindly searching for safety platforms. It is only the occasional safety position of the span; the rats in each treatment group have a clear search purpose, the swimming route is mainly concentrated in the safety platform quadrant, and the number of positions across the safety platform is significantly more than the model. Group.

 5 Conclusion

 The experiment showed that the spatial localization memory ability of the model group was seriously damaged. The spatial fixation memory ability of each treatment group was significantly improved compared with the model group, which was equivalent to the positive drug treatment. It was proved that the tryptophan derivatives of the present invention have a certain therapeutic effect on QA-induced AD rats. Example 11: Effect on lens in cataract rats

 1. Reagents and instruments:

Sodium selenite crystals (Na ₂ Se0 ₃ . 53⁄40 ) and Fuming tablets (Xi'an Beilin Pharmaceutical Co., Ltd.). Malondialdehyde (MDA) and superoxide dismutase (SOD) test kits (Nanjing Institute of Bioengineering). 756MC UV-Vis Spectrophotometer (Shanghai Precision Scientific Instrument Co., Ltd.). Digital display constant temperature three water tank (Jintou Guorui Experimental Instrument Factory, Jintan City, Jiangsu Province). Micropipette (Shanghai Qijing Biochemical Reagent Instrument Co., Ltd.). YZ-5CSI type slit lamp microscope (Suzhou Medical Instrument Factory).

 2. Animals:

 Healthy pure Wiatar rat suckling rats 100, both male and female, 10 days old, individual quality 14 ~ 16g, provided by the Experimental Animal Center of China Medical University.

 3. Grouping and cataract model establishment:

 Wistar rats were randomly divided into 5 groups: control group, model group, and tryptamine derivative group (Examples 1, 2, and 3, 50 mg/kg each), 20 in each group. Feed with breast milk and ordinary pellets, and drink freely. To the model group and the first group, the second group, and the third group, the rats were injected subcutaneously with a small dose (3.46 mg/kg body weight) of sodium selenite, once every other day for 3 times. Rats in the control group did not receive any treatment. The lens changes were observed with a slit lamp microscope on the third day after the last administration, and the lens opacity was successfully modeled.

 4. Administration and test methods:

 Each administration group was administered at a dose of 1 time/day for 2 weeks; the rats in the control group and the model group were given no drugs and were given free drinking water.

 (1) General observation of lens morphology: After the rats were molded, the mice were dilated with Meridian eye drops every week, and the rat lens was examined with a slit lamp microscope and photographed. The degree of opacity of the lens is divided into 0 ~ V period: the phase 0 is the lens transparent; the stage I is the lens around the cortex scattered in the small vacuole; the second stage is the lens surrounding cortex into a ring dense medium vacuole; the third stage is in addition to the lens surrounding cortical dense Outside the vacuoles, part of the cortex is turbid; in stage IV, the lens nucleus and perinuclear cortex are turbid; in stage V, the lens is completely turbid.

(2) Preparation of lens homogenate: Weigh the double-lens lens into a 5ml homogenized cup, add 8.6g/L ice physiological saline according to a certain ratio, and use a high-speed tissue masher to control the mash at 10000r/min. Min, 3.5% lens homogenate was prepared, then centrifuged at 2000 r/min for 8 min at low speed, and the supernatant was separated for MDA content and SOD activity. (3) Detection of MDA content and SOD activity in the lens: Take serum 100μ1, lens homogenate supernatant ΙΟΟμΙ, detect MDA content, take serum 30μ1, crystal homogenate supernatant 30μ1 to detect SOD activity, the specific process according to MDA and SOD test The box instructions are operated. Colorimetric measurements were performed using a Model 756MC UV-Vis spectrophotometer.

 5. Results

 The degree of opacity of the lens in each group of rats at the end of the experiment

 Group 0 Phase I Phase II Flood Stage IV Phase V Phase Control Group 2000 0 0 0 0 Model Group 0 0 0 4 11 3 Example 1 Group 3 10 6 1 0 0 Example 2 Group 2 11 5 2 0 0 Example 3 Group 3 9 7 1 0 0 During the experimental observation period, the lens of the control group was always transparent; in the model group, the surrounding cortex showed dense vacuoles and partial cortical opacity, which was III~V phase; The lens opacity of rats in groups 1, 2, and 3 was significantly reduced compared with the model group, which was ο~ πι. Table 5: Comparison of MDA content and SOD activity in rat serum and lens (X s) group lens (pair) MDA (nmol/mgprot) SOD (U/mgprot) control group 10 1.06±0.26 27.26±3.28 model group 10 1.51±0.27 16.64±3.96 Example 1 Group 10 1.23±0.30 20.17±3.58 Example 1 Group 10 1.18±0.26 22.54±2.87 Example 1 Group 10 1.24±0.18 21.36±3.15

The detection of MDA content showed that the MDA in the lens of the first, second and third groups was significantly lower than that of the model group, and the difference was significant (P<0.05). The detection of SOD activity showed that the SOD activity in the lens of the first, second and third groups was significantly higher than that of the model group, and the difference was significant (P<0.05).

 6 Conclusion

 The tryptophan derivatives of the present invention can alleviate the degree of lens opacity, which may be related to increasing SOD in the lens and lowering the MDA content, thereby counteracting and alleviating peroxidative damage. Example 12: Antidepressant effect on a mouse depression model

 Experimental animal

Male SPF Kunming mice, individual quality 20~25g, purchased from China Medical University Center of things.

 2. Animal grouping

 The mice were subjected to neurobehavioral testing and screening before the experimental grouping. The mice with different differences were screened out, and 40 mice were randomly selected and randomly divided into 4 groups, namely blank group. , tryptamine derivatives (Examples 1, 2, 3, 60 mg/kg per group), 10 per group.

 3. Test method:

 (1) Tail Suspension Test (TST)

 The mice in the 4 groups were acclimated to the environment for 1 week. The animals were kept in single cages 24 hours before the experiment, and fasting could not be stopped. On the day of the test, each group was intragastrically administered according to the dosage requirements. After lh administration, the tail of the mouse was glued to the tail-tailed balance scaffold at a distance of 2 cm from the tip of the tail, so that the tail of the mouse was not twisted and folded, and the head was turned to the head. The lower suspension is inverted, the head is 15cm away from the tabletop, each mouse is hovering for 6min, the first 2min is adapted, and the cumulative immobility time within 4min after the recording is 3⁄4. All the limbs are not moved except for breathing. .

 (2) Forced Swimming Test (FST)

 The mice in the 4 groups were acclimated to the environment for 1 week. The animals were trained to swim for 15 minutes before the experiment, and they were kept in a single cage. During the experiment, the mice in each group were dosed for 1 h, and placed in a circular container with a diameter of about 18 cm, a water depth of 18 cm, and a water temperature of (25 ± 1) °C. Accumulated immobility time (floating state, only the nostrils are exposed to keep breathing, and the limbs are occasionally swiped to keep the body from sinking).

 4. Results

 Table 6

 No time /s

 Grouping agent f (mg/kg)

 Suspended forced swimming blank group - 79.10±30.13 128.60±23.70 Example 1 group 60 52.30±20.47 82.34±26.37 Example 2 group 60 49.72±24.39 86.97±24.89 Example 3 group 60 54.81±23.10 77.15±22.81 The results are shown in Table 6. As shown, compared with the blank control group, the tryptamine derivative group (Examples 1, 2, and 3) all shortened the immobility time of swimming and tail suspension in mice, and there was no difference between the three groups.

 5 Conclusion

 The test results indicate that the tryptophan derivatives of the present invention are resistant to depressive symptoms caused by forced swimming and tail suspension in mice. Example 13: Effect on anxiety behavior in mice

Instrument Combined Open Field Test Analysis System, produced by Shanghai Jiliang Software Technology Co., Ltd.

 2. Animals and grouping

 Forty male mice in Kunming, with an individual mass (20 ± 2 ) g, were purchased from the Experimental Animal Center of China Medical University. After 1 week of adaptive feeding, the mice were randomly divided into 4 groups, namely, the blank group and the tryptamine derivative group (Examples 1, 2, and 3, each group of 60 mg/kg), and the mice started joint operation after lh administration. Behavior detection.

 3. Test method

 The mice were placed in the middle of the joint opening experiment box containing a 40 cm 40 cm 16-hole plate while timing was started. Infrared emitters and camera devices are placed in the joint opening experimental box, which can effectively record various spontaneous activity and caving behavior of mice. The depth of the hole is 2.2 cm, and infrared rays are placed at a depth of 1 cm. When the depth of the cavity of the mouse exceeds 1 cm, the instrument will automatically record the behavior of the cavity and accumulate the number of holes. Each mouse was recorded continuously for 6 minutes. The environment was kept quiet during the operation. The excretion in the box was cleaned before each time the mouse was placed, and the objectivity of each batch was ensured as much as possible.

 4. Results

 ^7

 Group number) Central area activity time (s) Central area activity distance (mm) Shuttle time (s) Blank group 10 37.65±27.00 249.50±149.96 33.20±15.56 Example 1 group 10 101.00±85.24 517.68±136.23 62.36±20.31 Implementation Example 2 Group 10 103.28±45.87 529.37±168.77 56.93±17.54 Example 3 Group 10 110.09±78.63 506.42士151.24 65.07士 25.91

1 Comparison of activity time in the central area: The tryptamine derivative group (Examples 1, 2, 3) was significantly different from the blank group (PO.05); 2 Comparison of activity distances in the central area: The tryptamine derivative group (Examples 1, 2, 3) were significantly different from the blank group (PO.05); 3 Shuttle time comparison: The tryptamine derivative group (Examples 1, 2, 3) compared with the blank group Significant difference (P < 0.05). Quantity) number of holes (times)

 Blank group 10 9.63±7.87 Example 1 Group 10 21.50±10.05 Example 2 Group 10 19.17±12.64 Example 3 Group 10 22.63±11.32

5 Conclusion:

In this joint opening animal self-issued test, the tryptophan derivative of the present invention was initially confirmed. The organism has an improved effect on the behavior of anxious mice. Example 14: Effect on adjuvant arthritis model rats

 Experimental animal

 SPF grade male Wistar rats aged 5-6 months, individual mass 180 ± 20g, provided by Experimental Animal Center, China Medical University.

 2. Reagents and instruments

 Liquid paraffin, produced by Tianjin Chemical Reagent Factory; medicinal lanolin, produced by Shanghai Huaheng Lanolin Factory; BCG for intradermal injection, produced by Shanghai Institute of Biological Products. 1/lOOmm electronic digital caliper, produced by Tianjin Measuring Tool Factory.

 3. Modeling and group administration

 (1) Preparation of Freund's complete adjuvant:

 After the anhydrous lanolin was heated and melted, 1 part was placed in a mortar, cooled slightly, and 2 parts of liquid paraffin was added while grinding. After grinding for 30 minutes, heat at 70 °C for 10 minutes, then autoclave for 1 hour, remove it, add 7.5 mg of BCG or inactivated tubercle bacillus per ml, grind evenly, store in 4 °C refrigerator, shake well before use.

 (2) Animal grouping and administration:

 After 50 days of adaptive feeding, 50 rats were randomly divided into 5 groups, namely blank group, model group and tryptophan derivative group (Examples 1, 2, 3, 50 mg/kg), 10 in each group. Animals in each group were intragastrically administered at 1 h before the onset of inflammation, and administered once a day after modeling for 3 days.

 (3) Preparation of adjuvant arthritis model:

 In addition to the normal control group, Freund's complete adjuvant (0.05 mL/only) was injected intradermally into the right hind paw of the rat. The normal control group was injected with an equal volume of normal saline.

 (4) Observation indicators:

 Swelling degree of the paws on the injection side (primary lesions): The thickness of the paws was measured by the caliper method before and after the inflammation, 18, 36, and 72 hours after the inflammation, and the degree of swelling was calculated.

 Swelling = thickness of the foot after injection - thickness of the foot before injection

 4. Results:

 Table 9 Effect on primary lesions in adjuvant arthritis model rats (n=10)

 Dose swelling degree (mm)

 Group

(mg/kg) 18h 36h 72h normal group - 0.01±0.05 0.01±0.06 0.01±0.04 model group - 2.94±0.57 3.30±0.41 3.21±0.34 Example 1 group 50 2.54±0.35 2.36±0.38* 2.19±0.31 * Example 2 groups 50 2.65±0.49 2.30±0.35* 2.15±0.43* Example 3 group 50 2.70±0.34 2.40±0.42* 2.20±0.25* Note: Compared with the model group, *P<0.05 or P< 0.01

 The results are shown in the above table. Compared with the model group, the tryptamine derivative group (Examples 1, 2, and 3) can significantly inhibit the paw swelling of the model side arthritis model rats after 36 hours of administration (PO). .05 or P< 0.01).

 5 Conclusion

 The results of the present experiment indicate that the intragastric administration of the tryptophan derivative of the present invention has a significant therapeutic effect on the primary lesion of the adjuvant arthritis model rat, and can suppress the immunological inflammation.

Claims

A compound of formula I or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof,

Wherein R ¹ or R ^{2 is} independently selected from the group consisting of hydrogen, C _r C ₅ alkyl, a substituent of the formula II, and one of R ¹ and R ² is a substituent of the formula II,

Wherein R ^{3 is} selected from aryl; n=0, 1, 2 or

2. The compound of claim 1 or a pharmaceutically acceptable salt, solvate, polymorph as claimed in claim enantiomer or racemic mixture, wherein, R ³ is a phenyl group; n = l.

The compound according to claim 1 or 2, or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof, characterized in that the other of R ¹ and R ² It is fluorine or argon.

The compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof, wherein R ¹ is fluorine or Hydrogen, R ² is a substituent of the formula II.

The compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof, wherein R ¹ is formula II The substituent shown, R ² is fluorine or hydrogen.

The compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof a solvate, a polymorph, an enantiomer or a racemic mixture, characterized in that the structure of the compound is as follows:

7. A process for the preparation of a compound according to any one of claims 1 to 6, or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof, which comprises the following step:

 In the presence of a base, a compound of the formula 与 is reacted with a compound of the formula XIV to form a compound of the formula I,

 XIII XIV.

 The preparation method according to claim 7, wherein the preparation method comprises the following steps:

(1) a compound of formula III is reacted with NaN ₃ in disulfoxide to form a compound of formula IV;

 III IV

 (2) a compound of the formula IV is reacted with a compound of the formula V in the presence of sodium ascorbate and cuprous iodide.

 V VI

(3) a compound of the formula VI is reacted with a reduced iron powder to form a compound of the formula VII

 Vii

 (4) The compound of formula VII is reacted with hydrated trichloroacetaldehyde and hydroxylamine hydrochloride in the presence of anhydrous sodium sulfate to form formula VIII.

 VIII

(5) reacting a compound of the formula VIII in a concentrated acid in the presence of sodium hydride to form a compound of the formula IX;

 IX

(6) reacting a compound of formula IX with a compound of formula X in the presence of triethylamine to form a compound of formula I,

Wherein, in Formula I, R ^{1 is} selected from the group consisting of hydrogen, CC ₅ alkyl, and halogen, and R ² is a substituent of Formula II;

 (7) In the presence of chromium trioxide, the compound of formula VIII is reacted in glacial acetic acid and acetic anhydride to form a compound of formula XI.

 XI

 (8) reacting a compound of the formula XI with a compound of the formula XII in the presence of triethylamine to form a compound of the formula I,

Wherein, in Formula I, R ¹ is a substituent of the formula II, and R ^{2 is} selected from the group consisting of hydrogen, C _r C ₅ alkyl and halogen.

The preparation method according to claim 7 or 8, wherein the preparation method comprises the following steps:

(1) The compound of the formula III is reacted with NaN ₃ at room temperature for 0.5-1 hour at room temperature. After completion of the reaction, water is added, and the mixture is extracted with ethyl acetate. The ethyl acetate phase is washed with water and dried over anhydrous sodium sulfate. An ester, that is, a compound of the formula IV;

 (2) The compound of the formula IV is reacted with sodium ascorbate, cuprous iodide, acetonitrile, water and a compound of the formula V at room temperature under a nitrogen atmosphere. After the reaction is completed, the reaction product is poured into water, and ethyl acetate is used. The extract is washed with a saturated aqueous solution of sodium chloride, and the ethyl acetate phase is dried over anhydrous sodium sulfate to remove ethyl acetate.

(3) The compound of the formula VI is reacted in an aqueous solution of ammonium chloride at 80 ° C for 4-5 hours in the presence of activated reduced iron powder. After the reaction, it is cooled to room temperature, and sodium carbonate is added to adjust the pH to 8-9. After adding ethyl acetate and stirring for 0.5 hours, the mixture was filtered through Celite, and the filtrate was extracted with ethyl acetate and washed with saturated brine to dryness, and ethyl acetate was dried over anhydrous sodium sulfate. Compound (4) The compound of the formula VII is stirred and mixed with hydrated trichloroacetaldehyde, anhydrous sodium sulfate and water, and then hydrochloric acid solution, hydroxylamine hydrochloride and water are sequentially added, and the reaction is carried out at 100 ° C for 2-3 hours. Cooling to room temperature, suction filtration, washing the cake with 10% by weight aqueous hydrochloric acid and water, and drying in vacuo to give the compound of formula VIII;

 (5) The compound of the formula VIII is dissolved in concentrated sulfuric acid at normal temperature and vigorous stirring, and then the temperature is raised to 65 ° C for 4-5 hours. After the reaction is completed, the reaction liquid is poured into cold water to precipitate a solid, and suction filtration is carried out. The filter cake is washed with water and dried under vacuum to obtain a compound of the formula IX;

 (6) The compound of the formula IX is reacted with the compound of the formula XII in terpene at 110 ° C for 3-4 hours in the presence of triethylamine. After completion of the reaction, the triethylamine and toluene are removed, and anhydrous ethanol is added. Recrystallization, that is, the compound of formula I,

Wherein, in Formula I, R ^{1 is} selected from the group consisting of hydrogen, CC ₅ alkyl, and halogen, and R ² is a substituent of Formula II;

 (7) In the presence of chromium trioxide, the compound of formula VIII is reacted in glacial acetic acid and acetic anhydride at 80-90 ° C for 3 hours. After the reaction is completed, the reaction product is cooled to room temperature, added with water, suction filtered, washed with water. Solid, suction filtration to obtain the compound of formula XI;

 (8) The compound of the formula XI is reacted with the compound of the formula XII in terpene at 110 ° C for 3 hours in the presence of triethylamine. After the reaction, the triethylamine and toluene are removed, and then ethanol is recrystallized. That is, the compound of formula I,

Wherein, in Formula I, R ¹ is a substituent of the formula II, and R ^{2 is} selected from the group consisting of hydrogen, C _r C ₅ alkyl and halogen.

10. The preparation method according to claim 8 or 9, wherein

In the step (1), the molar ratio of the compound of the formula III to NaN ₃ is 1:

1.2;

 In the step (2), the molar ratio of the compound of the formula IV to the sodium ascorbate, cuprous iodide, phenylacetylene is 1: 0.4: 0.2: 2;

 In the step (3), the molar ratio between the compound represented by the formula VI and the reduced iron powder is 1:8, and the concentration of the aqueous ammonium chloride solution is 5% by weight;

 In the step (4), the molar ratio of the compound of the formula VII to the hydrated trichloroacetaldehyde, sodium anhydrous sodium chloride, and hydroxylamine hydrochloride is 1: 1: 1:3, the hydrochloric acid The concentration of the aqueous solution is 5% by weight;

In the step (6), the molar ratio of the compound of the formula IX to the compound of the formula XII, triethylamine is 1: 1: 5; In the step (7), the molar ratio between the chromium trioxide and the compound of the formula VIII is 1: 1.1-1.2, and the molar ratio between the glacial acetic acid and acetic anhydride is 1:1. ;

 In the step (7), the molar ratio of the compound of the formula XI to the compound of the formula XII and triethylamine is 1 : 1 : 4-6.

The use of a compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof for the preparation of an IDO inhibitor .

The compound according to any one of claims 1 to 6, or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof, for the preparation and prevention or treatment of Amino-2,3-dioxygenase-mediated drug use in diseases associated with disorders of tryptophan metabolism,

 Preferably, the disease associated with a guanamine-2,3-dioxygenase-mediated disorder of tryptophan metabolism is selected from the group consisting of a tumor, a cancer, an Alzheimer's disease, an autoimmune disease, a cataract, a mental disorder. , one or more of depression and anxiety.

A pharmaceutical composition for a guanamine-2,3-dioxygenase inhibitor, the pharmaceutical composition comprising the compound according to any one of claims 1 to 6 or a pharmaceutically acceptable thereof Salts, solvates, polymorphs, enantiomers or racemic mixtures, and pharmaceutically acceptable carriers and/or excipients.

14. A method for treating, preventing or delaying a disease associated with a guanamine-2,3-dioxygenase mediated disorder of tryptophan metabolism, the method comprising administering to a patient in need thereof therapeutically effective A compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt, solvate, polymorph, enantiomer or racemic mixture thereof or a pharmaceutical combination according to claim 13 Object,

 Preferably, the disease associated with a guanamine-2,3-dioxygenase-mediated disorder of tryptophan metabolism is selected from the group consisting of a tumor, a cancer, an Alzheimer's disease, an autoimmune disease, a cataract, a mental disorder. , one or more of depression and anxiety.