CN111689952A - 2,3-dioxygenase inhibitor containing thioquaternary ring structure and preparation method and application thereof - Google Patents

Abstract

其中，

。The invention belongs to the field of pharmaceutical synthesis, and relates to a 1,2,5-oxadiazole compound containing a thioquaternary ring represented by the general formula (I) and a pharmaceutically acceptable salt thereof, a preparation method thereof, and a medical Applications. The compound of the present invention, its stereoisomer or its pharmaceutically acceptable salt or its pharmaceutical combination is combined with anti-CTLA-4 antibody, anti-PD-1 antibody, anti-PD-L1 antibody, antiviral agent, chemotherapeutic agent, immunosuppressive agent Indoleamine 2,3-dioxygenase activity, can be used for the treatment or prevention of cancer or tumor, virus Infections, depression, neurodegenerative disorders, trauma, age-related cataracts, organ transplant rejection or autoimmune disease.

in,

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Description

2,3-dioxygenase inhibitor containing thioquaternary ring structure and preparation method thereof use

technical field

The invention belongs to the field of drug synthesis and relates to 2,3-dioxygenase inhibitors, in particular to a 2,3-dioxygenase inhibitor containing a thio-quaternary ring structure and a preparation method and application thereof.

Background technique

Indoleamine-2,3-dioxygenase (IDO) is a monomeric enzyme containing heme that was first discovered in cells by Hayaishi's group in 1967. The cDNA encodes a protein composed of It is composed of 403 amino acids and has a molecular weight of 45kDa. It is widely distributed in human and other mammalian tissues other than the liver, such as lung, small and large intestine, rectum, spleen, kidney, stomach and brain, etc. It is the only one other than liver that can catalyze tryptophan. A rate-limiting enzyme for catabolism along the kynuric acid pathway. In the cells of tumor patients, IDO often plays an important physiological role in inducing immune tolerance in the tumor microenvironment, and its mediated tryptophan (Trp)-kynurenine (Kyn) metabolic pathway is involved in tumors Immune escape, and IDO also plays an important role in inducing immune tolerance in the tumor microenvironment.

Studies have shown that IDO can suppress local T cell immune responses in the tumor microenvironment through tryptophan depletion, toxic metabolism, and induction of regulatory T cell proliferation. In many cases, it is overexpressed in tumors, thereby consuming local tryptophan and producing a large amount of metabolites such as kynurenine. In fact, in the absence of tryptophan or kynurenine, T cells undergo proliferation inhibition, decreased activity and even apoptosis. There is a regulatory point in T cells that is very sensitive to the level of tryptophan. Under the action of IDO, tryptophan can be consumed, resulting in the arrest of T cells in the mid-G1 phase, thereby inhibiting the proliferation of T cells and inhibiting the T cell immune response. Once T cells stop proliferating, they may not be stimulated again, which is the immune mechanism of IDO in vivo. IDO has been confirmed to be closely related to Alzheimer's disease, cataract, cancer and other major human diseases, so IDO inhibitors can be used to treat cancer, Alzheimer's disease and other drugs with IDO-mediated tryptophan metabolic pathway Pathologically characterized diseases are receiving increasing attention as important drugs.

At present, the research and development of indoleamine 2,3-dioxygenase inhibitors are in the early stage, including Indoximod from NewLink, NLG-919, Epacadostat from Incyte (INCB 024360), and BMS, Flexus, Iomet, Iteos, Curadev and other companies' IDO or TDO inhibitors.

Based on the current state of the art, the inventors of the present application intend to provide a new IDO inhibitor, which will have important practical significance for improving the economic burden of tumor patients in my country and improving the clinical treatment effect of tumors.

SUMMARY OF THE INVENTION

The object of the present invention is to provide a new 2,3-dioxygenase inhibitor containing a thio-quaternary ring structure based on the current state of the prior art, and specifically relates to a 1,2,5-containing thio-quaternary ring structure - Potent IDO enzyme inhibitors of oxadiazoles.

Another object of the present invention is to provide a method for preparing the 1,2,5-oxadiazole compound containing a thio-quaternary ring.

Among the inhibitors of the present invention, the 1,2,5-oxadiazole compounds containing a thioquaternary ring and their pharmaceutically acceptable salts have the structural formula shown in the general formula (I),

in

In the present invention, preferred compounds have the following structures of compounds 1, 2, 3, and 4:

In the present invention, taking compound 3 as an example, the preparation process of the compound of the present invention is as follows:

In the formula, CDI is N,N'-carbonyldiimidazole; AC is amine carbamate, and DIB is iodobenzene acetate.

The present invention provides the use of a compound capable of inhibiting IDO, wherein the compound is used in the preparation of a pharmaceutical combination for preventing and/or treating a disease with pathological characteristics of IDO-mediated tryptophan metabolic pathway.

The compounds of the present invention can be added with pharmaceutically acceptable carriers to prepare common pharmaceutical preparations, such as tablets, capsules, powders, syrups, liquids, suspension injections and the like. The clinical administration mode of the compound of the present invention can be oral, injection and the like.

Another aspect of the present invention provides a method for modulating indoleamine 2,3-dioxygenase activity, the method comprising a therapeutically effective dose of the aforementioned compound, a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a medicament thereof Combination with anti-CTLA-4 antibody, anti-PD-1 antibody, anti-PD-L1 antibody, antiviral agent, chemotherapeutic agent, immunosuppressant, radiation, antitumor vaccine, antiviral vaccine, cytokine therapy or tyrosine kinase inhibition drug in combination.

The medicament of the present invention is used for treating or preventing cancer or tumor, viral infection, depression, neurodegenerative disorder, trauma, age-related cataract, organ transplant rejection or autoimmune disease; preferably, wherein the cancer or tumor is selected from lung cancer , Bone cancer, stomach cancer, pancreatic cancer, skin cancer, head and neck cancer, uterine cancer, ovarian cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, rectal cancer, colon cancer , Anal cancer, breast cancer, esophageal cancer, etc.

Detailed ways

Example 1: Synthesis of compound 1, N-(3-bromo-4-fluorophenyl)-N'-hydroxy-4-(1-oxathietan-3-yl)amino-1,2,5 -Oxadiazole-3-carboxamidine

1) Synthesis of 4-amino-N'-hydroxy-1,2,5-oxadiazole-3-carbaoxamidine

Malononitrile (0.5mol, 32g) was added to 1.5L of water preheated to 45°C, and after stirring for 5 minutes, sodium nitrite (0.55mol, 38g) was added under ice bath conditions. When the temperature reaches 10°C, add 5.5 ml of 6N hydrochloric acid, maintain the temperature at 16°C and stir for 15 minutes, remove the ice bath, and stir at 16-18°C for 1.5h. Cool down to 13°C. Dissolve 1.5mol, 104.2g of solid hydroxylamine hydrochloride in an appropriate amount of water, add an equal amount of sodium hydroxide to neutralize, add to the above reaction solution at one time, the temperature rises to 26 ° C, when the exothermic reaction is complete, remove the ice bath, 26 ~ 27 °C stirred for 1 h, then slowly heated to reflux for 2 h. After refluxing, it was cooled overnight. Post-processing: adjust to neutrality with aqueous sodium hydroxide solution, ice bath, suction filtration and dry to obtain 67.9 g of white solid product with a yield of 95%. ¹³ C NMR (100 MHz, CD ₃ OD): δ 156.0, 145.9, 141.3. MS ESI: m/z=144.0, [M+H] ⁺

2) Synthesis of 4-amino-N'-hydroxy-1,2,5-oxadiazole-3-carbaoxamidine chloride

Take 4-amino-N'-hydroxy-1,2,5-oxadiazole-3-carbaoxamidine (29.5mmol, 4.2g) and add it to a mixed solution of 59ml of water, 30ml of acetic acid and 6N hydrochloric acid, 42 Stir at ~45°C until complete dissolution. Add sodium chloride (88.5mmol, 5.2g), stir in an ice bath, dissolve 2g of sodium nitrite in water and slowly add it dropwise with a constant pressure dropping funnel, the temperature is maintained at 0 ℃, after the addition is completed, the ice bath is stirred for 1.5h and then heated to Stir at room temperature for 2h. Post-processing: suction filtration, washing with water, and drying to obtain 2.4 g of the product with a yield of 50%. ¹³ CNMR (100 MHz, CD ₃ OD): δ 155.8, 143.4, 129.7. MS ESI: m/z=162.9, [M+H] ⁺

3) Synthesis of 4-amino-N-(3-bromo-4-fluorophenyl)-N'-hydroxy-1,2,5-oxadiazole-3-carboxamidine

Dissolve 4-amino-N-hydroxy-1,2,5-oxadiazole-3 carbon heteroimide acid chloride (20mmol, 3.25g) in 45ml of water, heat to 60°C, 8.4g of 3-bromo-4- Fluoroaniline was added to the above solution and stirred for 10min, 2.5g of sodium bicarbonate was dissolved in hot water and added slowly, and stirred at 60°C for 20min. Suction filtration, washing with water, and drying to obtain 5.7 g of product with a yield of 90%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ 11.48 (s, J=3.1 Hz, 1H), 8.90 (s, 1H), 7.19 (t, J=8.7 Hz, 1H), 7.10 (dd, J= 6.1,2.7Hz,1H),6.77(dd,J=8.1,4.2Hz,1H),6.28(s,2H).MS ESI:m/z=317.1,[M+H] ⁺

4) Synthesis of 3-(4-amino-1,2,5-oxadiazol-3-yl)-4-(3-bromo-4-fluorophenyl)-1,2,4-oxadiazole-5 (4H)-keto

Dissolve 4-amino-N-(3-bromo-4-fluorophenyl)-N'-hydroxy-1,2,5-oxadiazole-3-carboxamidine (8.45mmol, 2.67g) in 20ml of tetrahydrofuran, 5.34g of N,N'-carbonyldiimidazole was dissolved in tetrahydrofuran and added to the above solution, heated to reflux for 1h, cooled to room temperature, rotary evaporated until a large amount of solid appeared, diluted with 50ml of ethyl acetate, washed twice with 30ml of 1N hydrochloric acid, Wash once with 30 ml of water, once with 30 ml of saturated brine, collect the organic layer, dry over anhydrous sodium sulfate, filter with suction, and spin dry to obtain 2.75 g of the product with a yield of 95%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ 7.19 (t, J=8.7Hz, 1H), 7.10 (dd, J=6.1, 2.7Hz, 1H), 6.77 (dd, J=8.1, 4.2Hz, 1H),6.28(s,2H).MS ESI: m/z＝343.0,[M+H] ⁺

5) Synthesis of 4-(3-bromo-4-fluorophenyl)-3-(4-nitro-1,2,5-oxadiazol-3-yl)-1,2,4-oxadiazole- 5(4H)-ketone

Take 3-(4-amino-1,2,5-oxadiazol-3-yl)-4-(3-bromo-4-fluorophenyl)-1,2,4-oxadiazole-5(4H )-ketone (10g, 29.2.0mmol) was added to 200ml of trifluoroacetic acid, 100ml of hydrogen peroxide was added, and reacted at 60°C for 12h. After the reaction, the reaction solution was poured into ice water, filtered and dried to obtain 7.6g of the product (brown yellow solid) , the yield is 70%. ¹ H NMR (400MHz, DMSO-d ₆ )δ8.09-8.01(m,1H),7.72-7.64(m,1H),7.62-7.53(m,1H).MS ESI:m/z=371.9,[ M+H] ⁺

6) Synthesis of 4-(3-bromo-4-fluorophenyl)-3-[4-(thietan-3-yl)amino)]-1,2,5-oxadiazol-3-yl )-1,2,4-Diazol-5(4H)-one

Take 3-(4-Amino-1,2,5-oxadiazol-3-yl)-4-(3-bromo-4-fluorophenyl)-1,2,4-oxadiazole-(4H) - Ketone (1.63 g, 4.4 mmol) was dissolved in tetrahydrofuran, 365 mg of 3-thietanamine was added to the above solution, stirred at room temperature for 2 h, and monitored by spot plate. Post-processing: spin off part of tetrahydrofuran, extract with dichloromethane, wash the organic phase with water, wash with saturated brine, dry with anhydrous sodium sulfate, use petroleum ether:ethyl acetate=12:1 as the eluent, column chromatography to obtain 1.04g of white Solid, 63% yield. ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.63 (dd, J=5.7, 2.6 Hz, 1H), 7.41-7.29 (m, 2H), 5.74 (d, J=7.7 Hz, 1H), 5.15-4.90 (m ,1H),3.53–3.46(m,4H).MS ESI:m/z＝414,[M+H] ⁺

7) Synthesis of 4-(3-bromo-4-fluorophenyl)-3-[4-[(1-oxathietane-3-yl)amino]-1,2,5-oxadiazole- 3-yl]-1,2,4-oxadiazol-5(4H)-one

Take 4-(3-bromo-4-fluorophenyl)-3-[4-(thietan-3-ylamino)-1,2,5-oxadiazol-3-yl]-1, 2,4-oxadiazol-5(4H)-one (200mg, 0.64mmol) was added to an eggplant flask with 10ml of dichloromethane and 1ml of water, and 214mg of 2-iodoylbenzoic acid (IBX) and a catalytic amount of benzyl Triethylammonium chloride (TEBA) was added to the above reaction solution and stirred at room temperature for 2h. Post-processing: extraction with dichloromethane, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using petroleum ether:ethyl acetate=10:1 as the eluent, and column chromatography to obtain 206 mg of product with a yield of 75%. ¹ H NMR (400MHz, CDCl ₃ )δ7.64(s,1H),7.39-7.29(m,2H),5.72(s,1H),5.11(s,1H),3.84-3.66(m,2H), 3.61–3.44(m,2H).MS ESI: m/z=430,[M+H] ⁺

8) Synthesis of N-(3-bromo-4-fluorophenyl)-N'-hydroxy-4-[(1-oxathietane-3-yl)amino]-1,2,5-

oxadiazole-3-carboxamidine

Take 4-(3-bromo-4-fluorophenyl)-3-[4-[(1-oxethitan-3-yl)amino]-1,2,5-oxadiazole-3- base]-1,2,4-oxadiazol-5(4H)-one (206 mg, 0.48 mmol) in tetrahydrofuran solution, 2.5 ml of 2N aqueous sodium hydroxide solution was added, and the reaction was carried out for 0.5 h. Post-processing: extraction with ethyl acetate, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using dichloromethane:methanol=30:1 as the eluent, and column chromatography to obtain 118 mg of product with a yield of 61%. ¹ H NMR (400MHz, DMSO-d ₆ )δ11.48(s, J=3.1Hz, 1H), 8.90(s, 1H), 7.33-7.08(m, 2H), 6.99(d, J=5.5Hz, 1H), 6.76(d, J＝8.8Hz, 1H), 4.59(dd, J＝13.9, 8.3Hz, 2H), 4.30(d, J＝7.8Hz, 1H), 4.21(dt, J＝14.5, 5.0 Hz, 2H). MS ESI: m/z=404, [M+H] ⁺ .

Example 2: Synthesis of compound 2, N-(3-bromo-4fluorophenyl)-N'-hydroxy-4-[(1,1'-dioxetan-3-yl)amino]- 1,2,5-oxadiazole-3-carboxamidine

1) Synthesis of 4-(3-Bromo-4-fluorophenyl)-3-(4-[(1,1′-dioxetan-3-yl)amino)-1,2,5- Oxadiazol-3-yl]-1,2,4-oxadiazol-5(4H)-one

Take 4-(3-bromo-4-fluorophenyl)-3-[4-(thietan-3-ylamino)-1,2,5-oxadiazol-3-yl]-1, 2,4-oxadiazol-5(4H)-one (200mg, 0.64mmol) was added to an eggplant-shaped flask with 10ml of dichloromethane, and 330mg of m-chloroperoxybenzoic acid was added to the reaction solution, and stirred at room temperature for 2h. Post-processing: extraction with dichloromethane, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using dichloromethane:methanol=50:1 as the eluent, and column chromatography to obtain 151 mg of product with a yield of 53%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ 7.33-7.08 (m, 2H), 6.96 (d, J=5.5Hz, 1H), 6.73 (d, J=8.8Hz, 1H), 4.59 (dd, J=13.9, 8.3Hz, 2H), 4.30 (d, J=7.8Hz, 1H), 4.23 (dt, J=14.5, 5.0Hz, 2H). MS ESI: m/z=446, [M+H] ⁺ 2) Synthesis of N-(3-bromo-4fluorophenyl)-N'-hydroxy-4-[(1,1'-dioxetan-3-yl)amino]-1,2, 5-oxadiazole-3-carboxamidine

Take 4-(3-bromo-4-fluorophenyl)-3-[4-[(1,1′-dioxetan-3-yl)amino]-1,2,5-oxanedi Azol-3-yl]-1,2,4-oxadiazol-5(4H)-one (151 mg, 0.34 mmol) was dissolved in tetrahydrofuran solution, 2.5 ml of 2N aqueous sodium hydroxide solution was added, and the reaction was carried out for 1 h. Post-processing: extraction with ethyl acetate, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using dichloromethane:methanol=30:1 as the eluent, and column chromatography to obtain 81 mg of product with a yield of 57%. ¹ H NMR (400MHz, DMSO-d ₆ )δ11.74-11.33(m,1H),8.98(s,1H),7.33-7.08(m,2H),6.99(d,J=5.5Hz,1H), 6.76(d,J＝8.8Hz,1H),4.59(dd,J＝13.9,8.3Hz,2H),4.30(d,J＝7.8Hz,1H),4.21(dt,J＝14.5,5.0Hz,2H) ).MS ESI: m/z=420, [M+H] ⁺ .

Example 3: Synthesis of compound 3, N-(3-bromo-4fluorophenyl)-N'-hydroxy-4-[(1-imino-1-oxo-1λ ⁶ thietane-3-yl ) Amino]-1,2,5-oxadiazole-3-carboxamidine

1) Synthesis of 4-(3-bromo-4-fluorophenyl)-3-[4-[(1-imino-1-oxo-1λ ⁶ thietane-3-yl)amino]-1, 2,5-oxadiazol-3-yl]-1,2,4-oxadiazol-5(4H)-one

Take 4-(3-bromo-4-fluorophenyl)-3-[4-(thietan-3-ylamino)-1,2,5-oxadiazol-3-yl]-1, 2,4-oxadiazol-5(4H)-one (500mg, 1.6mmol) was dissolved in methanol, 783mg of amine carbamate (AC) and 3.22g of iodobenzene acetate (DIB) were added to the above solution, stirred at room temperature, TLC monitoring. Post-processing: extraction with ethyl acetate, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using dichloromethane:methanol=50:1 as the eluent, and column chromatography to obtain 497 mg of product with a yield of 70%. ¹ H NMR (400MHz, Chloroform-d) δ 7.64 (dq, J=4.9, 2.4Hz, 1H), 7.39-7.32 (m, 1H), 7.29 (dd, J=7.5, 2.6Hz, 1H), 6.27 –6.17(m,1H),4.67–4.42(m,2H),4.15–3.94(m,2H).MSESI:m/z＝445,[M+H]+

2) Synthesis of N-(3-bromo-4 fluorophenyl)-N'-hydroxy-4-[(1-imino-1-oxo-1λ ⁶ thietane-3-yl)amino]-1 ,2,5-oxadiazole-3-carboxamidine

Take 4-(3-bromo-4-fluorophenyl)-3-[4-[(1-imino-1-oxo-1λ ⁶ thietane-3-yl)amino]-1,2, 5-oxadiazol-3-yl]-1,2,4-oxadiazol-5(4H)-one (494 mg, 1.12 mmol) was dissolved in tetrahydrofuran, 4 ml of 2N aqueous sodium hydroxide solution was added, and the reaction was carried out for 1 h. Post-processing: extraction with ethyl acetate, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using dichloromethane:methanol=30:1 as the eluent, and column chromatography to obtain 248 mg of product with a yield of 53%. ¹ H NMR (400 MHz, CDCl ₃ ) δ 11.61 (d, J=3.0 Hz, 1H), 8.96 (s, 1H), 7.20 (qd, J=8.5, 3.1 Hz, 1H), 7.12 (dq, J= 6.1, 2.7Hz, 1H), 6.88 (d, J=6.5Hz, 1H), 6.78 (dp, J=7.6, 4.0, 3.3Hz, 1H), 4.71 (d, J=7.4Hz, 1H), 4.49– 4.24 (m, 3H), 3.96 (tt, J=8.5, 4.3 Hz, 2H). MS ESI: m/z=419, [M+H] ⁺ .

Example 4: Synthesis of compound 4, N-(3-bromo-4fluorophenyl)-4-[[1-(cyanoimino)-1λ ⁴ -thietan-3-yl]amino]- N'-Hydroxy-1,2,5-oxadiazole-3-carboxamidine

1) Synthesis of N-[3-[[4-(3-bromo-4fluorophenyl)-5-oxo-4,5dihydro-1,2,4-oxadiazol-3-yl]-1 ,2,5-thioxadiazol-3-yl]amino]-1λ ⁴ -thietane-1-ylidene)cyanamide

Take 4-(3-bromo-4-fluorophenyl)-3-[4-(thietan-3-ylamino)-1,2,5-oxadiazol-3-yl]-1, 2,4-oxadiazol-5(4H)-one (200 mg, 0.64 mmol) and 53.76 mg were dissolved in acetonitrile, 309 mg of iodobenzene acetate was added under an ice bath, the ice bath was removed, and the mixture was stirred at room temperature for 3 h. Post-processing: extraction with ethyl acetate, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using dichloromethane:methanol=50:1 as the eluent, and column chromatography to obtain 217 mg of product with a yield of 73%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ 8.10(s, 1H), 7.73(s, 1H), 7.66–7.47(m, 2H), 4.40(s, 1H), 4.25(s, 2H), 4.02(t,J＝10.5Hz,2H).MS ESI:m/z＝454,[M+H] ⁺

2) Synthesis of N-(3-bromo-4-fluorophenyl)-4-[[1-(cyanoimino)-1λ ⁴ -thietan-3-yl]amino]-N'-hydroxy- 1,2,5-oxadiazole-3-carboxamidine

Take N-[3-[[[4-(3-bromo-4fluorophenyl)-5-oxo-4,5dihydro-1,2,4-oxadiazol-3-yl]-1, A solution of 2,5-thioxadiazol-3-yl]amino]-1λ ⁴ -thietan-1-ylidene]cyanamide (217mg, 0.47mmol) in tetrahydrofuran was added 4ml of 2N aqueous lithium hydroxide solution , the reaction is 1h. Post-processing: extraction with ethyl acetate, washing the organic phase with water, washing with saturated brine, drying over anhydrous sodium sulfate, using dichloromethane:methanol=30:1 as the eluent, and column chromatography to obtain 112 mg of product with a yield of 56%. ¹ HNMR (400MHz, DMSO-d ₆ )δ11.51(s, 1H), 8.99(s, 1H), 7.18(t, J=8.7Hz, 1H), 7.14-7.10(m, 1H), 7.06(d , J=6.1Hz, 1H), 6.74 (dt, J=7.9, 3.1Hz, 1H), 4.94 (q, J=6.9, 6.4Hz, 1H), 4.00 (td, J=14.7, 12.9, 6.6Hz, 4H). MS ESI: m/z=428, [M+H] ⁺ .

Example 5: Enzymatic evaluation of IDO activity inhibition

Buffer (pH=6.5) preparation: take 599.9 mg of sodium dihydrogen phosphate, add 1.52 mL of 1 M potassium hydroxide, add water to dilute to 100 mL, and then fine-tune the pH to 6.5. Preparation of standard reaction solution: using buffer solution (pH=6.5) as mother solution, adding 176.12 mg of ascorbic acid, 10 mg of catalase, 0.325 mg of methylene blue, and 4.075 mg of L-tryptophan. All compounds to be tested were prepared into 80 mmol DMSO solution, and then 80 mmol was diluted into 8 μmol with Buffer for use. Take a 96-well black plate, add 80 μL of standard reaction solution to test wells, maximum and minimum control wells; add 10 μL of IDO enzyme and 10 μL of test compound solution to test wells, and add 10 μL of enzyme and Add 10 μL of Buffer, and add 20 μL of Buffer to the smallest control wells. After standing at 37°C for 1 hour, take it out, add 20 μl of 1M sodium hydroxide solution to the wells of plate 96, and place it at 60°C for 4 hours (the purpose is to decompose N-formylkynurenine into kynurenine) After taking out, after cooling, use a microplate reader to test the fluorescence value. Inhibition rate=(maximum value-measured value)/(maximum value-minimum value)*100%.

The results (as shown in Table 1) showed that the compounds of the present invention showed better IDO enzymatic activity, wherein the IC ₅₀ values of compounds 2 and 4 for IDO enzymatic activity were less than 100 nM. The compounds of the present invention can be further developed to develop IDO inhibitors.

Table 1: IDO enzyme inhibitory activity IC ₅₀ values of the compounds of the present invention

compound IDO enzyme inhibitory activity IC₅₀(nM) Compound 1 212.40±46.83 Compound 2 98.85±6.13 Compound 3 207.97±9.96 Compound 4 69.21±6.76

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Example 6: Cell model test of IDO activity inhibition

The model that interferon gamma induces the expression of IDO in Hela cells was used to test the inhibitory activity of compounds against indoleamine 2,3-dioxygenase (IDO). The compound to be tested was prepared with dimethyl sulfoxide at 8 mM, diluted with dimethyl sulfoxide to the highest concentration in the experiment, and the medium was used for 3-fold gradient dilution during the experiment, generally diluted to 7 to 10 concentration points, each concentration was set. Duplicate holes. The final concentration of dimethyl sulfoxide was 0.5%, and the internal reference compound was included in each experiment;

The experimental procedure was as follows: in vitro cultured HeLa cells were seeded in a 96-well plate (10 ⁴ cells/well, 100 μL) (ie, 10 ⁵ cells/ml, 100 μl per well), and after culturing for 24 h, human IFN-γ (80 ng /mL), gradient dilution inhibitor, L-tryptophan (15 μg/mL), D++ in total 200 μL (ie, the total volume in the final well 200 μl) was added to the 96-well plate and mixed. After culturing for 48 h, centrifuge at 1200 rpm for 3 min, take 140 μL of the supernatant into another 96-well plate, add 10 μL of 6.1 M trichloroacetic acid to mix, and incubate at 50°C for 30 min. Then centrifuge at 2500rpm for 10min, take 100μL of the supernatant into another 96-well plate, add 100μL of 2% (w/v) 4-(dimethylamino)benzaldehyde (acetic acid solution) to react until the solution turns yellow, and then place it in a 96-well plate. Read at 480nm wavelength. Inhibition rate %={(MAX-A)*100/(MAX-MIN) where MAX is the reading without inhibitor, MIN is the reading without interferon and without inhibitor (all parts without adding are filled with D++ ), A is the reading of adding inhibitor;

The results are shown in Table 2. The results show that the compounds of the present invention show better IDO enzymatic activity in cells, and the IC ₅₀ values of compounds 1-4 for IDO enzymatic activity in Hela cells are all less than 100 nM. The compounds of the present invention can be further developed to develop IDO inhibitors.

Table 2: _IC50 values of IDO inhibitory activity in cells of the compounds of the present invention

compound IDO activity inhibition IC₅₀(nM) in Hela cells Compound 1 78.81±15.62 Compound 2 59.46±7.22 Compound 3 53.27±11.47 Compound 4 72.67±16.03

.

Claims (11)

1. An indoleamine-2,3-dioxygenase inhibitor containing a sulfonamide structure is characterized in that the inhibitor contains a 1,2, 5-oxadiazole compound of a thio four-membered ring shown in a general formula (I) and pharmaceutically acceptable salts thereof,

wherein,

2. the indoleamine-2,3-dioxygenase inhibitor containing a sulfonamide structure according to claim 1, wherein the 1,2, 5-oxadiazole compound having a thiofour-membered ring is compound 1,

3. the indoleamine-2,3-dioxygenase inhibitor containing a sulfonamide structure according to claim 1, wherein the 1,2, 5-oxadiazole compound having a thiofour-membered ring is compound 2 having the following structure,

4. the indoleamine-2,3-dioxygenase inhibitor containing a sulfonamide structure according to claim 1, wherein the 1,2, 5-oxadiazole compound having a thiofour-membered ring is compound 3,

5. the indoleamine-2,3-dioxygenase inhibitor containing a sulfonamide structure according to claim 1, wherein the 1,2, 5-oxadiazole compound having a thiofour-membered ring is compound 4,

6. the process for preparing indoleamine-2,3-dioxygenase inhibitor containing a sulfonamide structure according to claim 1, wherein the process for preparing the same is represented by the following formula,

wherein CDI is N, N' -carbonyldiimidazole; AC is carbamate amine and DIB is iodobenzene acetate.

7. Use of the indoleamine 2,3-dioxygenase inhibitor containing a sulfonamide structure according to claim 1 for the preparation of a pharmaceutical composition for the prophylaxis and/or treatment of diseases with pathological features of the IDO-mediated tryptophan metabolic pathway.

8. The use according to claim 7, wherein the disease is: cancer or tumor, viral infection, depression, neurodegenerative disorder, trauma, age-related cataract, organ transplant rejection or autoimmune disease.

9. Use according to claim 8, wherein the cancer or tumour is selected from lung cancer, bone cancer, stomach cancer, pancreatic cancer, skin cancer, head and neck cancer, uterine cancer, ovarian cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, rectal cancer, colon cancer, cancer of the anal region, breast cancer or oesophageal cancer.

10. A pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (i) as claimed in claim 1 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

11. The pharmaceutical composition of claim 10, wherein the pharmaceutical composition is administered in combination with an anti-CTLA-4 antibody, an anti-PD-1 antibody, an anti-PD-L1 antibody, an antiviral agent, a chemotherapeutic agent, an immunosuppressive agent, radiation, an anti-tumor vaccine, an antiviral vaccine, cytokine therapy, or a tyrosine kinase inhibitor.