A diarylpyrimidine derivative containing a fused heterocycle and a preparation method and application thereof

Abstract

The application discloses a kind of diaromatic pyrimidine compounds containing fused heterocycle and preparation method and application thereof.The compound has the structure shown in general formula I.The application further relates to pharmaceutical compositions containing the compound of formula I.Active screening experiments show that the compound of the application has good anti-HIV and CHIKV activity, and therefore the application further provides the use of the above-mentioned compound in the preparation of anti-AIDS drugs and anti-Chikungunya virus drugs.

Description

TECHNICAL FIELD

[0001] The present application relates to a derivative and a preparation method and application thereof, in particular to a diarylpyrimidine derivative containing a fused heterocycle and a preparation method and application thereof, and belongs to the technical field of medicines. BACKGROUND

[0002] Acquired immune deficiency syndrome (AIDS) is a serious infectious disease that destroys the human immune system mainly caused by human immunodeficiency virus type 1 (HIV-1) infection. In the life cycle of HIV-1, reverse transcriptase (RT) is responsible for reverse transcription of single-stranded RNA carrying viral genetic information into double-stranded DNA, and is an optimal target for anti-AIDS drug design. As an important component of combined antiretroviral therapy (cART), non-nucleoside reverse transcriptase inhibitors (NNRTIs) inhibit the activity of HIV-1 RT through allosteric binding, have the advantages of high activity, strong selectivity, low toxicity, etc., and are a hot research direction of anti-AIDS drugs.

[0003] Etravirine (ETR) and rilpivirne (RPV) are new generation diarylpyrimidine NNRTIs. However, with their extensive use in the clinic, a variety of mutant strains resistant to ETR and RPV have emerged, such as E138A, E138K, Y181C and Y181I, etc. In addition, this type of compound has low water solubility and oral bioavailability, large oral dose, and is prone to cause more serious side effects in clinical use. Therefore, it is one of the important research fields of anti-AIDS drugs to obtain NNRTIs with high efficiency, broad-spectrum resistance and good pharmacokinetic properties through reasonable structural modification.

[0004] Chikungunya fever (CHIKF) is a mosquito-borne infectious disease caused by Chikungunya virus (CHIKV) and widely spread in the world. Clinically, it is characterized by fever, rash, bleeding, and joint pain. A small number of patients are left with persistent joint stiffness. Chikungunya fever is prone to large-scale epidemic and outbreak in areas with high mosquito density, and is one of the mosquito-borne infectious diseases that seriously affect human public health safety in the early 21st century. At present, there is no vaccine or antiviral drug available for the prevention and control of CHIKV infection, so it is urgent to develop safe and effective small molecule drugs with clinical application value to treat Chikungunya fever. SUMMARY

[0005] The application provides a kind of containing fused heterocycle diaryl pyrimidine derivative and preparation method thereof, and the application also provides the partial activity screening result of the above-mentioned compound and its use.

[0006] The technical scheme of the application is as follows:

[0007] I. containing fused heterocycle diaryl pyrimidine derivative

[0008] The containing fused heterocycle diaryl pyrimidine derivative of the application has the structure shown in the following general formula I:

[0009]

[0010] Among them,

[0011] R1 is CH3, CN, p-C4H6-CN or CH=CHCN;

[0012] R2 is H, CH3, CN, CF3, COCH3, NH2, NO2, F, Br, I, Cl, PO(CH3)2, PO(OCH3)2, SO2CH3, SOCH3, OCH3, SCH3, NHCH3, NHCOCH3 or NHSO2CH3;

[0013] R3 is H, CH3, CN, CF3, COCH3, NH2, NO2, F, Br, I, Cl, PO(CH3)2, PO(OCH3)2, SO2CH3, SOCH3, OCH3, SCH3, NHCH3, NHCOCH3 or NHSO2CH3;

[0014] X is

[0015]

[0016] According to the application, the containing fused heterocycle diaryl pyrimidine derivative is one of the following:

[0017]

[0018]

[0019] II. Preparation method of diarylpyrimidine derivatives containing fused heterocycle

[0020] The preparation method of the diarylpyrimidine derivatives containing fused heterocycle comprises the following steps: first, using 2,4-dichloropyrimidine 1 as a raw material, an intermediate 2 is generated by reacting with (E)-3,5-dimethyl-4-hydroxyphenylacrylonitrile in an N,N-dimethylformamide solution; the intermediate 2 is coupled with an aniline of the fused heterocycle under the catalysis of palladium acetate to obtain a target compound 3(a-x) or an intermediate 4 or 7; then, the intermediate 4 or 7 is removed from a Boc group to obtain an intermediate 5 or 8, and the intermediate 5 or 8 is reacted with different sulfonyl chlorides, phosphoryl chlorides, acyl chlorides or halogenated alkanes to obtain final products 6(a-e) or 9(a-e);

[0021]

[0022] Reagents and conditions: (i) (E)-3,5-dimethyl-4-hydroxyphenylacrylonitrile, N,N-dimethylformamide, potassium carbonate, 50 DEG C; (ii) palladium acetate, 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene, cesium carbonate, nitrogen, 1,4-dioxane, 90 DEG C; (iii) trifluoroacetic acid, dichloromethane, room temperature; (iv) sulfonyl chloride, phosphoryl chloride, acyl chloride or halogenated alkane, triethylamine, 0 DEG C.

[0023] III. Application of diarylpyrimidine derivatives containing fused heterocycle

[0024] The present application discloses the anti-HIV activity screening results and anti-CHIKV results of the diarylpyrimidine compounds containing fused heterocycle, and the first application of the diarylpyrimidine compounds as HIV inhibitors and CHIKV inhibitors. The experiments prove that the diarylpyrimidine compounds containing fused heterocycle provided by the present application can be used for preparing anti-HIV drugs and anti-CHIKV drugs. The present application also provides the application of the above-mentioned compounds in preparing anti-HIV drugs and anti-CHIKV drugs.

[0025] Therefore, the diarylpyrimidine compounds containing fused heterocycle provided by the present application can be used as HIV inhibitors and CHIKV inhibitors, and can be used for preparing anti-HIV drugs and anti-CHIKV drugs.

[0026] A pharmaceutical composition for resisting HIV and CHIKV contains the above-mentioned diarylpyrimidine compounds containing fused heterocycle and pharmaceutically acceptable salts thereof and pharmaceutical adjuvants, and is prepared into different dosage forms. DETAILED DESCRIPTION

[0027] The present application will be further understood by the following examples, which are not intended to limit the scope of the present application.

[0028] Example 1: Preparation of (E)-3-(4-((2-chloropyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (2)

[0029]

[0030] To a solution of 2,4-dichloropyrimidine (1.49 g, 0.01 mol) and potassium carbonate (1.66 g, 0.012 mol) in N,N-dimethylformamide (20 mL), (E)-3,5-dimethyl-4- hydroxyphenylacrylonitrile (1.73 g, 0.01 mol) was added and stirred at 50 °C for 4 h until the reaction was complete. Ice water (200 mL) was added and extracted with ethyl acetate (3 x 50 mL). The combined organic layers were washed with saturated brine, dried over anhydrous Na2S04, filtered and concentrated under reduced pressure and finally recrystallized using ethyl acetate and petroleum ether to afford the pure intermediate 4-((2-chloropyrimidin-4-yl)oxy)-3,5-dimethylbenzonitrile 2. White solid, yield: 84.7%. 1 HNMR (400 MHz, DMSO-d6) δ 8.67 (d, J = 5.7 Hz, 1H, pyrimidine-H), 7.62 (d, J = 16.7 Hz, 1H, CH=), 7.52 (s, 2H, Ph-H), 7.25 (d, J = 5.7 Hz, 1H, pyrimidine-H), 6.45 (d, J = 16.7 Hz, 1H, CH=), 2.07 (s, 6H, Ph-CH3x2). ESI-MS: m / z 286.3 (M+H) + ,C 15 H 12 ClN3O (285.07).

[0031] Example 2: Preparation of (E)-3-(4-((2-(benzo[d]thiazol-5-ylamino)pyrimidin-4- yl)oxy)-3,5-dimethylphenyl)acrylonitrile (3a)

[0032]

[0033] (E)-3-(4-((2-chloropyrimidin-4-yl)oxy)-3,5-dimethylphenyl)acrylonitrile (0.52 g, 3.01 mmol), benzo[d]thiazol-5-amine (0.45 g, 3.01 mmol), palladium acetate (0.034 g, 0.15 mmol), 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (0.086 g, 0.15 mmol) and cesium carbonate (1.46 g, 4.51 mmol) were dissolved in 20 mL of 1,4-dioxane, protected by nitrogen, and reacted at 90 °C for 8 hours. The reaction completion was monitored by TLC. After cooling to room temperature, the mixture was filtered and the filtrate was concentrated under reduced pressure. Further purification was performed by silica gel chromatography with methanol / dichloromethane as eluent to obtain the target compound 3a. Brown solid, yield: 47.4%. 1 HNMR (600 MHz, DMSO-d6) δ 9.81 (s, 1H, NH), 9.24 (s, 1H, thienyl-H), 8.44 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.30 (s, 1H, Ph-H), 7.82 (d, J = 8.5 Hz, 1H, Ph-H), 7.64 (d, J = 16.7 Hz, 1H, CH=), 7.57 (d, J = 8.8 Hz, 1H, Ph-H), 7.52 (s, 2H, Ph-H), 6.54 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.44 (d, J = 16.7 Hz, 1H, CH=), 2.12 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.78, 160.70, 160.27, 156.56, 154.23, 152.49, 150.67, 139.72, 134.04, 131.58, 129.85, 128.87, 125.87, 121.98, 120.15, 118.47, 112.51, 98.68, 96.71, 17.53. ESI-MS: m / z 400.08 (M+H) + ,C 22 H 17 N5OS (399.12).

[0034] Example 3: Preparation of (E)-3-(3,5-dimethyl-4-((2-(2-methylbenzo[d]thiazol-5-yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (3b)

[0035]

[0036] Preparation method is same with example 2, only change raw material to 2-methylbenzo[d]thiazol-5-amine. Yellow solid, yield: 45.4%. 1 H NMR (600 MHz, DMSO-d6) δ 9.77 (s, 1H, NH), 8.42 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.04 (s, 1H, Ph-H), 7.68 (s, 1H, Ph-H), 7.65 (d, J = 16.7 Hz, 1H, CH=), 7.52 (s, 2H, Ph-H), 7.41 (s, 1H, Ph-H), 6.55 (d, J = 7.2 Hz, 1H, pyrimidyl-H), 6.44 (d, J = 16.7 Hz, 1H, CH=), 2.75 (s, 3H, CH3), 2.11 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.78, 167.31, 160.72, 160.23, 154.13, 152.09, 150.91, 139.11, 131.92, 131.49, 128.90, 128.17, 121.45, 119.47, 117.46, 111.71, 97.95, 96.51, 20.25, 16.56. ESI-MS: m / z 414.28 (M+H) + ,C 23 H 19 N5OS (413.13).

[0037] Example 4: (E)-3-(4-((2-(benzo[d]thiazol-6-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (3c)

[0038]

[0039] Preparation method is same with example 2, only change raw material to 2-methylbenzo[d]thiazol-5-amine. Yellow solid, yield: 45.4%. 1H NMR (600 MHz, DMSO-d6) δ 9.99 (s, 1H, NH), 9.19 (s, 1H, thienyl-H), 8.46 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.05 (s, 1H, Ph-H), 7.82 (d, J = 8.9 Hz, 1H, Ph-H), 7.72 (d, J = 16.7 Hz, 1H, CH=), 7.59 (s, 2H, Ph-H), 7.43 (d, J = 9.0 Hz, 1H, Ph-H), 6.65 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.51 (d, J = 16.7 Hz, 1H, CH=), 2.12 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.76, 160.93, 160.02, 153.75, 151.96, 150.66, 148.67, 148.14, 138.61, 134.72, 131.81, 129.76, 128.76, 122.97, 118.96, 109.60, 98.17, 96.96, 16.56. ESI-MS: m / z 400.04 (M+H) + ,C 22 H 17 N5OS (399.12).

[0040] Example 5: (E)-3-(4-((2-(1,3-dihydroisobenzofuran-5-yl)amino)pyrimidin-4-yl)oxy)-3,5-dimethylphenyl)acrylonitrile

[0041]

[0042] Preparation method is same as example 2, only change the raw material to 1,3-dihydroisobenzofuran-5-amine. Brown solid, yield: 48.3%. 1 H NMR (600 MHz, DMSO-d6) δ 9.73 (s, 1H, NH), 8.39 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.68 (d, J = 16.7 Hz, 1H, CH=), 7.54 (s, 2H, Ph-H), 7.31 (s, 1H, Ph-H), 7.14 (s, 1H, Ph-H), 7.01 (d, J = 8.1 Hz, 1H, Ph-H), 6.56 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.49 (d, J = 16.7 Hz, 1H, CH=), 4.86 (s, 2H, OCH2), 4.63 (s, 2H, OCH2), 2.09 (s, 6H). 13C NMR (100 MHz, DMSO-d6) δ 168.71, 160.81, 160.17, 152.39, 150.52, 140.03, 139.70, 131.85, 131.71, 129.63, 128.63, 121.19, 119.38, 118.24, 110.54, 97.64, 96.78, 73.35, 72.78, 16.65. ESI-MS: m / z 385.03 (M+H) + ,C 23 H 20 N4O2 (384.16).

[0043] Example 6: (E)-3-(3,5-dimethyl-4-((2-((3-oxoisoindolin-5-yl)amino)pyrimidin-4- yl)oxy)phenyl)acrylonitrile (3e)

[0044]

[0045] Preparation method is same as example 2, only change the raw material to 6-aminoisoindolin-1-one. Grey solid, yield 44.2%. 1 HNMR (600 MHz, DMSO-d6) δ 9.78 (s, 1H, NH), 8.47 (s, 1H, amide-H), 8.42 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.91 (s, 1H, Ph-H), 7.67 (d, J = 7.3 Hz, 1H, Ph-H), 7.63 (d, J = 16.7 Hz, 1H, CH=), 7.51 (s, 2H, Ph-H), 7.21 (s, 1H, Ph-H), 6.52 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 6.44 (d, J = 16.7 Hz, 1H, CH=), 4.25 (s, 2H, CH2), 2.12 (d, J = 15.1 Hz, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 170.50, 168.73, 160.67, 160.19, 152.01, 150.57, 140.42, 137.26, 133.37, 131.75, 131.66, 128.83, 123.68, 122.36, 119.45, 113.01, 98.24, 96.77, 44.89, 16.48. ESI-MS: m / z 396.07 (M-H) - ,C 23 H 19 N5O2 (397.15).

[0046] Example 7: (E)-3-(3,5-dimethyl-4-(2-(2-methyl-1,3-dioxoisoindolin-5-yl)amino)pyrimidin-4- yl)oxy)phenyl)acrylonitrile (3f)

[0047]

[0048] Preparation method is same as example 2, only change raw material to 5-amino-2-methylisoindole-1,3-dione. Yellow solid, yield 50.7%. 1 H NMR (600 MHz, DMSO-d6) δ 10.25 (s, 1H, NH), 8.51 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 8.07 (s, 1H, Ph-H), 7.81 (d, J = 8.1 Hz, 1H, Ph-H), 7.64 (d, J = 16.7 Hz, 1H, CH=), 7.53 (s, 2H, Ph-H), 7.50 (d, J = 8.3 Hz, 1H, Ph-H), 6.68 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.45 (d, J = 16.7 Hz, 1H, CH=), 2.98 (s, 3H, CH3), 2.11 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.83, 168.28, 168.16, 160.81, 159.67, 151.92, 150.49, 146.25, 133.68, 131.90, 131.59, 128.88, 123.94, 123.88, 122.39, 119.33, 112.77, 99.70, 96.84, 24.05, 16.52. ESI-MS: m / z 424.12 (M-H) - ,C 24 H 19 N5O3 (425.15).

[0049] Example 8: (E)-3-(4-((2-(benzo[d][1,3]dioxol-5-ylamino)pyrimidin-4-yl)oxy)-3,5-dimethylphenyl)acrylonitrile (3g)

[0050]

[0051] Preparation method is same as example 2, only change raw material to benzo[d][1,3]dioxol-5-amine. Brown solid, yield 52.8%. 1H NMR (600 MHz, DMSO-d6) δ 9.47 (s, 1H, NH), 8.34 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.62 (d, J = 16.7 Hz, 1H, CH=), 7.49 (s, 2H, Ph-H), 7.04 (s, 1H, Ph-H), 6.82 (s, 1H, Ph-H), 6.60 (s, 1H, Ph-H), 6.46 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.42 (d, J = 16.7 Hz, 1H, CH=), 5.88 (s, 2H, CH2), 2.08 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.67, 160.67, 160.16, 152.19, 150.62, 147.33, 142.04, 135.08, 131.82, 131.65, 128.70, 119.39, 111.66, 107.94, 101.43, 101.07, 97.41, 96.60, 16.53. ESI-MS: m / z 387.05 (M+H) + ,C 22 H 18 N4O3 (386.14).

[0052] Example 9: (E)-3-(3,5-dimethyl-4-(2-(2-methylbenzo[d]oxazol-5-yl)amino)pyrimidin-4- yl)oxy)phenyl)acrylonitrile (3h)

[0053]

[0054] Preparation method is same as example 2, only change the raw material to 2-methylbenzo[d]oxazol-5-amine. Yellow solid, yield 47.7%. 1 H NMR (600 MHz, DMSO-d6) δ 9.47 (s, 1H, NH), 8.34 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.62 (d, J = 16.7 Hz, 1H, CH=), 7.49 (s, 2H, Ph-H), 7.04 (s, 1H, Ph-H), 6.82 (s, 1H, Ph-H), 6.60 (s, 1H, Ph-H), 6.46 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.42 (d, J = 16.7 Hz, 1H, CH=), 5.88 (s, 2H, CH2), 2.08 (s, 6H, CH3x2). 13C NMR (100 MHz, DMSO-d6) δ 168.73, 164.42, 160.73, 160.25, 152.12, 150.84, 145.99, 141.72, 137.26, 131.94, 131.60, 128.83, 119.48, 116.62, 109.79, 109.38, 97.71, 96.56, 16.18, 14.55. ESI-MS: m / z 398.12 (M+H) + ,C 23 H 19 N5O2 (387.15).

[0055] Example 10: (E)-3-(4-((2-(benzofuran-5-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (3i)

[0056]

[0057] Preparation method was the same as Example 2, only the raw material was changed to benzofuran-5-amine. Brown solid, yield 50.7%. 1 H NMR (600 MHz, DMSO-d6) δ 9.64 (s, 1H, NH), 8.38 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.89 (d, J = 2.2 Hz, 1H, furan-H), 7.72 (s, 1H, furan-H), 7.69 (d, J = 16.7 Hz, 1H, CH=), 7.57 (s, 2H, Ph-H), 7.34 - 7.19 (m, 3H, Ph-H), 6.53 (d, J = 4.9 Hz, 1H, pyrimidyl-H), 6.49 (d, J = 16.7 Hz, 1H, CH=), 2.11 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.73, 164.42, 160.73, 160.25, 152.12, 150.84, 145.99, 141.72, 137.26, 131.94, 131.60, 128.83, 119.48, 116.62, 109.79, 109.38, 97.71, 96.56, 16.18, 14.55. ESI-MS: m / z 398.12 (M+H) + ,C 23 H 18 N4O2 (382.14).

[0058] Example 11: (E)-3-(4-((2-(2,2-difluorobenzo[d][l,3]dioxol-5-yl)amino)pyrimidin-4- yl)oxy)-3,5-dimethylphenyl)acrylonitrile (3j)

[0059]

[0060] Preparation method is same as example 2, only change raw material to 2,2-difluorobenzo[d][l,3]dioxol-5-amine. Red solid, yield 45.2%. 1 H NMR (600 MHz, DMSO-d6) δ 9.88 (s, 1H, NH), 8.42 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.62 (d, J = 16.7 Hz, 1H, CH=), 7.53 (s, 2H, Ph-H), 7.33 (s, 1H, Ph-H), 7.13 (d, J = 8.7 Hz, 1H, Ph-H), 7.04 (s, 1H, Ph-H), 6.61 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.43 (d, J = 16.7 Hz, 1H, CH=), 2.08 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.70, 160.95, 159.79, 152.15, 150.48, 142.97, 137.53, 137.32, 132.02, 131.66, 128.70, 119.29, 114.04, 109.96, 101.08, 98.20, 96.63, 15.95. ESI-MS: m / z 421.07 (M-H) - ,C 22 H 16 F2N4O3 (422.12).

[0061] Example 12: (E)-3-(4-((2-([l,2,4]triazolo[4,3-a]pyridin-6-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (3k).

[0062]

[0063] Preparation method is same as example 2, only change raw material to [l,2,4]triazolo[4,3-a]pyridin-6-amine. White solid, yield 51.8%. 1H NMR (400 MHz, DMSO-d6) δ 9.99 (s, 1H, NH), 9.11 (s, 1H, triazole-H), 8.50 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.30 (s, 1H, Ph-H), 7.68 (s, 2H, Ph-H), 7.64 (d, J = 16.6 Hz, 1H, CH=), 7.53 (s, 2H, Ph-H), 6.64 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 6.43 (d, J = 16.6 Hz, 1H, CH=), 2.11 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.89, 160.80, 159.78, 153.71, 151.84, 150.67, 146.17, 132.13, 131.46, 129.90, 128.96, 125.85, 119.41, 117.06, 115.76, 98.55, 96.80, 16.53. ESI-MS: m / z 384.20 (M+H) + ,C 21 H 17 N7O (383.15).

[0064] Example 13: (E)-3-(4-((2-(benzothiophen-5-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (31)

[0065]

[0066] Preparation method is same as example 2, only change the raw material to benzothiophen-5-amine. White solid, yield 51.8%. 1 H NMR (400 MHz, DMSO-d6) δ 9.99 (s, 1H, NH), 9.11 (s, 1H, triazole-H), 8.50 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.30 (s, 1H, Ph-H), 7.68 (s, 2H, Ph-H), 7.64 (d, J = 16.6 Hz, 1H, CH=), 7.53 (s, 2H, Ph-H), 6.64 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 6.43 (d, J = 16.6 Hz, 1H, CH=), 2.11 (s, 6H, CH3x2). 13C NMR (100 MHz, DMSO-d6) δ 168.89, 160.80, 159.78, 153.71, 151.84, 150.67, 146.17, 132.13, 131.46, 129.90, 128.96, 125.85, 119.41, 117.06, 115.76, 98.55, 96.80, 16.53. ESI-MS: m / z 384.20 (M+H) + ,C 21 H 17 N7O(383.15)。

[0067] Example 14: (E)-3-(3,5-dimethyl-4-((2-(2-methyl-l-oxoisoindolin-5- yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (3m)

[0068]

[0069] Preparation method is same as example 2, only change the raw material to 5-amino-2- methylisoindolin-l-one. White solid, yield 42.9%. 1 H NMR (600 MHz, DMSO-d6) δ 10.07 (s, 1H, NH), 8.46 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.72 (d, J = 16.7 Hz, 1H, CH=), 7.58 (s, 2H, Ph-H), 7.53 (s, 1H, Ph-H), 7.38 (d, J = 8.3 Hz, 1H, Ph-H), 7.30 (d, J = 8.4 Hz, 1H, Ph-H), 6.67 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.55 (d, J = 16.7 Hz, 1H, CH=), 3.97 (s, 2H, -CH2), 3.00 (s, 3H, -N-CH3), 2.10 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.89, 160.80, 159.78, 153.71, 151.84, 150.67, 146.17, 132.13, 131.46, 129.90, 128.96, 125.85, 119.41, 117.06, 115.76, 98.55, 96.80, 16.53. ESI-MS: m / z 384.20 (M+H) + ,C 24 H 21 N5O2(411.17)。

[0070] Example 15: (E)-6-((4-(4-(2-cyanoethenyl)-2,6-dimethylphenoxy)pyrimidin-2-yl)amino)benzo[d]thiazole-2-carbonitrile (3n)

[0071]

[0072] Preparation method was same as example 2, only the raw material was changed to 6- aminobenzo[d]thiazole-2-carbonitrile. White solid, yield 42.9%. 1 H NMR (600 MHz, DMSO-d6) δ 10.34 (s, 1H, NH), 8.51 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.18 (s, 1H, Ph-H), 8.01 (d, J = 9.0 Hz, 1H, Ph-H), 7.78 (d, J = 16.7 Hz, 1H, CH=), 7.61 (s, 1H, Ph-H), 7.59 (s, 2H, Ph-H), 6.74 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.54 (d, J = 16.8 Hz, 1H, CH=), 2.11 (s, 6H, CH3x2). 13 C NMR (150 MHz, DMSO-d6) δ 168.76, 161.02, 159.70, 152.29, 150.60, 146.57, 141.69, 137.22, 133.03, 132.04, 131.88, 128.76, 124.82, 121.20, 119.28, 114.00, 108.33, 99.08, 97.14, 16.63. ESI-MS: m / z 425.09 (M+H) + ,C 23 H 16 N6OS (424.11).

[0073] Example 16: (E)-3-(4-((2-(benzo[d]isoxazol-5-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (3o)

[0074]

[0075] Preparation method was same as example 2, only the raw material was changed to benzo[d]isoxazol-5-amine. Brown solid, yield 48.7%. 1H NMR (400 MHz, DMSO-d6) δ 10.60 (s, 1H, NH), 9.59 (s, 1H, -CH=N-), 8.37 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.65 (s, 1H, Ph-H), 7.58 (d, J = 16.7 Hz, 1H, CH=), 7.50 (s, 2H, Ph-H), 7.42 (s, 1H, Ph-H), 6.73 (d, J = 8.5 Hz, 1H, Ph-H), 6.52 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.40 (d, J = 16.7 Hz, 1H, CH=), 2.09 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.75, 160.73, 160.05, 155.34, 152.02, 150.56, 132.79, 131.77, 131.54, 128.79, 126.20, 122.10, 119.44, 117.36, 116.61, 98.56, 97.93, 96.74, 16.53. ESI-MS: m / z 384.03 (M+H) + ,C 22 H 17 N5O2(383.14).

[0076] Example 17: (E)-3-(4-((2-((2,3-dihydrobenzo[b][l,4]dioxin-6-yl)amino)pyrimidin-4- yl)oxy)-3,5-dimethylphenyl)acrylonitrile (3p)

[0077]

[0078] Preparation method is same as example 2, only change the raw material to 2,3- dihydrobenzo[b][l,4]dioxin-6-amine. Brown solid, yield 49.7%. 1 H NMR (400 MHz, DMSO-d6) δ 10.60 (s, 1H, NH), 9.59 (s, 1H, -CH=N-), 8.37 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.65 (s, 1H, Ph-H), 7.58 (d, J = 16.7 Hz, 1H, CH=), 7.50 (s, 2H, Ph-H), 7.42 (s, 1H, Ph-H), 6.73 (d, J = 8.5 Hz, 1H, Ph-H), 6.52 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.40 (d, J = 16.7 Hz, 1H, CH=), 2.09 (s, 6H, CH3x2). 13C NMR (100 MHz, DMSO-d6) δ 168.75, 160.73, 160.05, 155.34, 152.02, 150.56, 132.79, 131.77, 131.54, 128.79, 126.20, 122.10, 119.44, 117.36, 116.61, 98.56, 97.93, 96.74, 16.53. ESI-MS: m / z 384.03 (M+H) + ,C 22 H 17 N5O2 (383.14).

[0079] Example 18: (E)-3-(4-((2-(benzo[d]isothiazol-6-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (3q)

[0080]

[0081] Preparation method was the same as example 2, only the raw material was changed to benzo[d]isothiazol-6-amine. Brown solid, yield 52.6%. 1 HNMR (600 MHz, DMSO-d6) δ 9.98 (s, 1H, NH), 8.53 (s, 1H, isothiazole-H), 8.46 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.18 (s, 1H, Ph-H), 7.94 (d, J = 8.8 Hz, 1H, Ph-H), 7.72 (d, J = 16.7 Hz, 1H, CH=), 7.61 (s, 2H, Ph-H), 7.56 (d, J = 9.1 Hz, 1H, Ph-H), 6.63 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.53 (d, J = 16.7 Hz, 1H, CH=), 2.13 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.77, 160.86, 160.20, 155.39, 152.33, 150.39, 144.72, 138.34, 136.97, 131.91, 131.82, 128.77, 121.85, 120.34, 119.35, 111.37, 98.22, 97.01, 15.95. ESI-MS: m / z 400.03 (M+H) + ,C 22 H 17 N5OS (399.12).

[0082] Example 19: (E)-3-(4-((2-(benzo[c][l,2,5]thiadiazol-5-ylamino)pyrimidin-4- yl)oxy)-3,5-dimethylphenyl)acrylonitrile (3r)

[0083]

[0084] Preparation method is same as example 2, only change the raw material to benzo[c][l,2,5]thiadiazol-5-amine. Brown solid, yield 50.4%. 1 H NMR (600 MHz, DMSO-d6) δ 10.16 (s, 1H, NH), 8.53 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 8.16 (s, 1H, Ph-H), 7.85 (d, J = 9.4 Hz, 1H, Ph-H), 7.74 (d, J = 7.4 Hz, 1H, Ph-H), 7.67 (d, J = 16.7 Hz, 1H, CH=), 7.55 (s, 2H, Ph-H), 6.69 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.45 (d, J = 16.7 Hz, 1H, CH=), 2.12 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.82, 160.83, 159.95, 156.00, 152.00, 151.7, 150.80, 141.78, 132.14, 131.50, 128.97, 126.79, 120.90, 119.44, 105.02, 99.28, 96.70, 16.55. ESI-MS: m / z 399.10 (M-H) - ,C 21 H 16 N6OS (400.11).

[0085] Example 20: (E)-3-(4-((2-(imidazo[l,2-a]pyridin-6-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (3s)

[0086]

[0087] Preparation method is same as example 2, only change the raw material to imidazo[l,2-a]pyridin-6-amine. White solid, yield 49.7%. 1H NMR (600 MHz, DMSO-d6) δ 9.78 (s, 1H, NH), 8.46 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.71 (d, J = 16.7 Hz, 1H, CH=), 7.61 (s, 2H, Ph-H), 7.44 (s, 2H, imidazole-H), 7.41 (s, 1H, Ph-H), 7.23 (d, J = 9.7 Hz, 2H), 6.63 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.52 (d, J = 16.7 Hz, 1H, CH=), 2.12 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.81, 160.88, 159.92, 152.20, 150.35, 142.04, 133.19, 131.93, 131.89, 129.76, 128.80, 127.99, 121.31, 119.34, 116.95, 113.82, 97.86, 97.12, 16.18. ESI-MS: m / z 383.21 (M+H) + ,C 22 H 18 N6O (382.15).

[0088] Example 21: (E)-3-(4-((2-(imidazo[l,2-a]pyridin-6-ylamino)pyrimidin-4-yl)oxy)-3,5- dimethylphenyl)acrylonitrile (3s)

[0089]

[0090] Preparation method is same as example 2, only change the raw material to imidazo[l,2-a]pyridin-6-amine. White solid, yield 49.7%. 1 H NMR (600 MHz, DMSO-d6) δ 9.78 (s, 1H, NH), 8.46 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.71 (d, J = 16.7 Hz, 1H, CH=), 7.61 (s, 2H, Ph-H), 7.44 (s, 2H, imidazole-H), 7.41 (s, 1H, Ph-H), 7.23 (d, J = 9.7 Hz, 2H), 6.63 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.52 (d, J = 16.7 Hz, 1H, CH=), 2.12 (s, 6H, CH3x2). 13C NMR (100 MHz, DMSO-d6) δ 168.81, 160.88, 159.92, 152.20, 150.35, 142.04, 133.19, 131.93, 131.89, 129.76, 128.80, 127.99, 121.31, 119.34, 116.95, 113.82, 97.86, 97.12, 16.18. ESI-MS: m / z 383.21 (M+H) + ,C 22 H 18 N6O (382.15).

[0091] Example 22: (E)-3-(3,5-dimethyl-4-((2-((l-methyl-lH-indazol-5-yl)amino)pyrimidin-4- yl)oxy)phenyl)acrylonitrile (3t)

[0092]

[0093] Preparation method was the same as Example 2, only the raw material was changed to 1- methyl-lH-indazol-5-amine. Pink solid, yield 48.2%. 1 H NMR (600 MHz, DMSO-d6) δ 9.66 (s, 1H, NH), 8.39 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.73 (d, J = 16.6 Hz, 2H, CH=; imidazole-H), 7.59 (s, 2H, Ph-H), 7.54 (s, 1H, Ph-H), 7.38 (d, J = 8.7 Hz, 1H, Ph-H), 7.33 (s, 1H, Ph-H), 6.54 (d, J = 2.2 Hz, 1H, pyrimidyl-H), 6.52 (d, J = 9.0 Hz, 1H, CH=), 3.96 (s, 3H, CH3), 2.11 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.72, 160.73, 160.37, 152.46, 150.58, 136.31, 133.82, 132.06, 131.84, 131.72, 128.73, 123.86, 120.89, 119.39, 109.67, 108.19, 97.27, 96.77, 35.73, 16.57. ESI-MS: m / z 397.14 (M+H) + ,C 23 H 20 N6O (396.17).

[0094] Example 23: (E)-3-(3,5-dimethyl-4-((2-(2-oxoindolin-6-yl)amino)pyrimidin-4- yl)oxy)phenyl)acrylonitrile (3u)

[0095]

[0096] Preparation method is same as example 2, only change raw material to 6-aminoindolin-2- one. Yellow solid, yield 51.4%. 1 H NMR (600 MHz, DMSO-d6) δ 10.21 (s, 1H, -CONH), 9.53 (s, 1H, NH), 8.36 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.63 (d, J = 16.7 Hz, 1H, CH=), 7.51 (s, 2H, Ph-H), 7.11 (s, 1H, Ph-H), 7.06 (d, J = 8.2 Hz, 1H, Ph-H), 6.81 (s, 1H, Ph-H), 6.46 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.43 (d, J = 16.7 Hz, 1H, CH=), 3.34 (s, 2H, CH2), 2.09 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 177.25, 168.68, 160.56, 160.27, 152.11, 150.45, 144.16, 140.06, 131.75, 129.68, 128.72, 124.30, 119.37, 119.00, 111.76, 101.13, 97.77, 96.82, 35.72, 16.18. ESI-MS: m / z 396.23 (M-H) - ,C 23 H 19 N5O2 (397.15).

[0097] Example 24: (E)-3-(3,5-dimethyl-4-((2-(4-methyl-2-oxo-2H-chromen-7-yl)amino)pyrimidin- 4-yl)oxy)phenyl)acrylonitrile (3v)

[0098]

[0099] Preparation method is same as example 2, only change raw material to 7-amino-4-methyl-2H- chromen-2-one. Grey solid, yield 47.8%. 1H NMR (600 MHz, DMSO-d6) δ 10.11 (s, 1H, NH), 8.48 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.64 (d, J = 16.7 Hz, 1H, CH=), 7.54 (s, 1H, Ph-H), 7.53 (s, 2H, Ph-H), 7.42 (d, J = 10.5 Hz, 1H, Ph-H), 7.32 (d, J = 7.8 Hz, 1H, Ph-H), 6.67 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.43 (d, J = 16.7 Hz, 1H, CH=), 6.17 (s, 1H, CH), 2.34 (s, 3H, CH3), 2.11 (s, 6H, CH3x2). ESI-MS: m / z 425.07 (M+H) + ,C 25 H 20 N4O3 (424.15).

[0100] Example 25: (E)-3-(3,5-dimethyl-4-((2-(2-oxo-4-(trifluoromethyl)-2H-chromen-7-yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (3w)

[0101]

[0102] Preparation method is same as example 2, only change the raw material to 7-amino-4- (trifluoromethyl)-2H-chromen-2-one. Brown solid, yield 46.2%. 1 H NMR (600 MHz, DMSO-d6) δ 10.11 (s, 1H, NH), 8.48 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.64 (d, J = 16.7 Hz, 1H, CH=), 7.54 (s, 1H, Ph-H), 7.53 (s, 2H, Ph-H), 7.42 (d, J = 10.5 Hz, 1H, Ph-H), 7.32 (d, J = 7.8 Hz, 1H, Ph-H), 6.67 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.43 (d, J = 16.7 Hz, 1H, CH=), 6.17 (s, 1H, CH), 2.34 (s, 3H, CH3), 2.11 (s, 6H, CH3x2). ESI-MS: m / z 425.07 (M+H) 13C NMR (100 MHz, DMSO-d6) δ 168.78, 160.96, 159.52, 159.20, 155.31, 152.03, 150.34, 145.21, 139.83 (dd, J = 32.1 Hz), 131.97, 131.60, 128.84, 124.99, 122.16 (dd, J = 275.5 Hz), 120.19, 119.32, 118.05, 113.22 (dd, J = 5.9 Hz), 106.88, 105.35, 99.67, 96.86, 17.00. ESI-MS: m / z 477.07 (M-H)-, C 25 H 17 F3N4O3 (478.13).

[0103] Example 26: (E)-3-(3,5-dimethyl-4-(2-((l-oxo-l,2,3,4-tetrahydroisoquinolin-7- yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (3x)

[0104]

[0105] Preparation method was the same as Example 2, only the starting material was changed to 7-amino-3,4-dihydroisoquinolin-l(2H)-one. White solid, yield 50.7%. 1 H NMR (400 MHz, DMSO-d6) δ 9.64 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.94 (s, 1H, -CONH), 7.85 (s, 1H, Ph-H), 7.64 (d, J = 16.4 Hz, 1H, CH=), 7.54 (s, 1H, Ph-H), 7.51 (s, 2H, Ph-H), 6.89 (d, J = 8.5 Hz, 1H, Ph-H), 6.48 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.44 (d, J = 16.8 Hz, 1H, CH=), 3.31 (d, J = 3.0 Hz, 2H, CH2), 2.77 (t, J = 6.6 Hz, 2H, CH2), 2.10 (s, 6H, CH3x2). 13H NMR (400 MHz, DMSO-d6) δ 9.64 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.94 (s, 1H, -CONH), 7.85 (s, 1H, Ph-H), 7.64 (d, J = 16.4 Hz, 1H, CH=), 7.54 (s, 1H, Ph-H), 7.51 (s, 2H, Ph-H), 6.89 (d, J = 8.5 Hz, 1H, Ph-H), 6.48 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.44 (d, J = 16.8 Hz, 1H, CH=), 3.31 (d, J = 3.0 Hz, 2H, CH2), 2.77 (t, J = 6.6 Hz, 2H, CH2), 2.10 (s, 6H, CH3x2). + H 24 H 21 N5O2 (411.17).

[0106] Example 27: (E)-3-(3,5-dimethyl-4-(2-((1-oxo-1,2,3,4-tetrahydroisoquinolin-7- yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (3y)

[0107]

[0108] Preparation method is same with example 2, only change the raw material to 7-amino-3,4- dihydroisoquinolin-1(2H)-one. White solid, yield 50.7%. 1 H NMR (400 MHz, DMSO-d6) δ 9.64 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.94 (s, 1H, -CONH), 7.85 (s, 1H, Ph-H), 7.64 (d, J = 16.4 Hz, 1H, CH=), 7.54 (s, 1H, Ph-H), 7.51 (s, 2H, Ph-H), 6.89 (d, J = 8.5 Hz, 1H, Ph-H), 6.48 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.44 (d, J = 16.8 Hz, 1H, CH=), 3.31 (d, J = 3.0 Hz, 2H, CH2), 2.77 (t, J = 6.6 Hz, 2H, CH2), 2.10 (s, 6H, CH3x2). 13 H NMR (400 MHz, DMSO-d6) δ 9.64 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.94 (s, 1H, -CONH), 7.85 (s, 1H, Ph-H), 7.64 (d, J = 16.4 Hz, 1H, CH=), 7.54 (s, 1H, Ph-H), 7.51 (s, 2H, Ph-H), 6.89 (d, J = 8.5 Hz, 1H, Ph-H), 6.48 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.44 (d, J = 16.8 Hz, 1H, CH=), 3.31 (d, J = 3.0 Hz, 2H, CH2), 2.77 (t, J = 6.6 Hz, 2H, CH2), 2.10 (s, 6H, CH3x2). + H 24 H21 N5O2 (411.17).

[0109] Example 28: (E)-7-((4-(4-(2-cyanoethenyl)-2-methylphenoxy)pyrimidin-2-yl)amino)- 3,4-dihydroisoquinoline-2(lH)-carboxylic acid tert-butyl ester (4)

[0110]

[0111] Preparation method was same as example 2, only the starting material was changed to 7-amino-3,4-dihydroisoquinoline-2(lH)-carboxylic acid tert-butyl ester. Brown solid, yield 44.3%. 1 H NMR (600 MHz, DMSO-d6) δ 9.58 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.62 (d, J = 16.7 Hz, 1H, CH=), 7.51 (s, 2H, Ph-H), 7.21 (s, 2H, Ph-H), 6.85 (s, 1H, Ph-H), 6.54 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.41 (d, J = 16.4 Hz, 1H, CH=), 4.08 (s, 2H, CH2), 3.48 (t, J = 5.9 Hz, 2H, CH2), 2.61 (t, J = 5.9 Hz, 2H, CH2), 2.11 (s, 6H, CH3x2), 1.46 (s, 9H, CH3x3). ESI-MS: m / z 498.40 (M+H) + ,C 29 H 31 N5O3 (497.24).

[0112] Example 29: (E)-3-(3,5-dimethyl-4-(2-(l,2,3,4-tetrahydroisoquinolin-7-yl)amino)pyrimidin- 4-yl)oxy)phenyl)acrylonitrile (5)

[0113]

[0114] Dissolve 1.0 equivalent of compound 4 in 15 mL of dichloromethane, add 3.0 equivalent of trifluoroacetic acid, stir at room temperature for 2 h, monitor the reaction completion by TLC. Adjust pH value to 9 with saturated sodium carbonate solution, dichloromethane extraction (3 x 10 mL), combine the organic phase, wash with saturated sodium chloride (3 x 20 mL), dry over anhydrous sodium sulfate, filter and concentrate the filtrate under reduced pressure. Recrystallize from ethyl acetate to get crude 5. White solid, yield 84.3%. ESI-MS: m / z 398.07 (M+H) + ,C 24 H 23N5O (397.19).

[0115] Example 30: (E)-7-(4-(4-(2-cyanoethenyl)-2,6-dimethylphenoxy)pyrimidin-2-yl)amino)- 3,4-dihydroisoquinoline-2(lH)-sulfonamide (6a)

[0116]

[0117] (E)-3-(3,5-dimethyl-4-(2-(l,2,3,4-tetrahydroisoquinolin-7-yl)amino)pyrimidin-4- yl)oxy)phenyl)acrylonitrile 5 (0.15 g, 0.37 mmol) was dissolved in 10 mL of dichloromethane, triethylamine (62 μL, 0.45 mmol) and sulfamoyl chloride (0.043 g, 0.37 mmol) were added slowly under ice bath condition, the reaction was monitored by TLC for 4 h at 0 °C. The reaction was quenched with water (20 mL) and extracted with dichloromethane (3 x 8 mL), the organic layers were combined and washed with saturated sodium chloride (3 x 15 mL), dried over anhydrous sodium sulfate, filtered and the filtrate was concentrated under reduced pressure, the compound 6a was recrystallized from ethyl acetate. White solid, yield 84.8 %. 1 HNMR (600 MHz, DMSO-d6) δ 9.53 (s, 1H, NH), 8.37 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.61 (d, J = 16.7 Hz, 1H, CH=), 7.51 (s, 2H, Ph-H), 7.26 (s, 1H, Ph-H), 7.18 (s, 1H, Ph-H), 6.89 (s, 2H, NH2), 6.83 (d, J = 8.3 Hz, 1H, Ph-H), 6.51 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.41 (d, J = 16.7 Hz, 1H, CH=), 3.97 (s, 2H, CH2), 3.20 (t, J = 5.8 Hz, 2H, CH2), 2.76 (t, J = 5.9 Hz, 2H, CH2), 2.11 (s, 6H, CH3x2). 13 CNMR (150 MHz, DMSO-d6) δ 168.75, 160.66, 160.28, 152.22, 150.39, 138.65, 132.70, 131.78, 131.72, 128.87, 128.68, 126.71, 119.36, 117.78, 116.21, 97.80, 96.87, 48.24, 44.33, 27.94, 17.40. ESI-MS: m / z 477.06 (M+H) + ,C 24 H 24N6O3S (476.16).

[0118] Example 31: (E)-3-(3,5-dimethyl-4-((2-(methylsulfonyl)-1,2,3,4-tetrahydroisoquinolin-7- yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (6b)

[0119]

[0120] Preparation method is the same as example 30, only the raw material is changed to methanesulfonyl chloride. White solid, yield 82.6%. 1 HNMR (600 MHz, DMSO-d6) δ 9.61 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.61 (d, J = 16.7 Hz, 1H, CH=), 7.52 (s, 2H, Ph-H), 7.25 (s, 1H, Ph-H), 7.16 (s, 1H, Ph-H), 6.88 (d, J = 8.4 Hz, 1H, Ph-H), 6.54 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.43 (d, J = 16.7 Hz, 1H, CH=), 4.02 (s, 2H, CH2), 3.34 (d, J = 5.7 Hz, 2H, -CH2), 2.92 (s, 3H, methylsulfonyl-CH3), 2.76 (t, J = 5.9 Hz, 2H, -CH2), 2.10 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.75, 160.69, 160.24, 152.22, 150.48, 138.80, 132.35, 131.75, 129.62, 129.13, 128.66, 126.56, 119.41, 117.94, 115.73, 97.81, 96.90, 47.52, 43.67, 35.33, 27.97, 16.59. ESI-MS: m / z 476.06 (M+H) + ,C 25 H 25 N5O3S (475.17).

[0121] Example 32: (E)-(7-((4-(4-(2-cyanoethenyl)-2,6-dimethylphenoxy)pyrimidin-2-yl)amino)- 3,4-dihydroisoquinolin-2(lH)-yl)dimethyl phosphonate (6c)

[0122]

[0123] The preparation method was the same as that of Example 30, except that the raw material was changed to dimethyl chlorophosphate. White solid, yield 83.7%. 1 H NMR (600 MHz, DMSO-d6) δ 9.57 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.64 (d, J = 16.7 Hz, 1H, CH=), 7.52 (s, 2H, Ph-H), 7.16 (s, 1H, Ph-H), 7.07 (s, 1H, Ph-H), 6.84 (d, J = 7.2 Hz, 1H, Ph-H), 6.54 (d, J = 5.4 Hz, 1H, pyrimidyl-H), 6.45 (d, J = 16.7 Hz, 1H, CH=), 3.84 (s, 2H, -CH2), 3.58 (s, 3H, -OCH3), 3.56 (s, 3H, -OCH3), 3.22 (m, 2H, -CH2), 2.61 (t, J = 6.0 Hz, 2H, -CH2), 2.10 (s, 6H, CH3x2). 13 C NMR (150 MHz, DMSO-d6) δ 168.76, 160.68, 160.26, 152.26, 150.47, 138.55, 131.81, 131.73, 129.60, 129.37, 128.66, 127.17, 119.41, 117.60, 115.29, 97.67, 96.86, 53.00, 46.51, 42.23, 27.99, 16.58. ESI-MS: m / z 506.42 (M+H) + ,C 26 H 28 N5O4P (505.19).

[0124] Example 33: (E)-(7-((4-(4-(2-cyanoethenyl)-2,6-dimethylphenoxy)pyrimidin-2-yl)amino)-3,4-dihydroisoquinolin-2(lH)-yl) dimethyl phosphonate (6d)

[0125]

[0126] The preparation method was the same as that of Example 30, except that the raw material was changed to 2-methylsulfonylacetyl chloride. White solid, yield 72.7%. 1H NMR (400 MHz, DMSO-d6) δ 9.62 (s, 1H, NH), 8.39 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.63 (d, J = 16.7 Hz, 1H, CH=), 7.54 (s, 2H, Ph-H), 7.25 (s, 1H, Ph-H), 7.12 (s, 1H, Ph-H), 6.87 (d, J = 8.4 Hz, 1H, Ph-H), 6.54 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.41 (d, J = 16.6 Hz, 1H, CH=), 4.55 (s, 2H, CH2), 4.31 (s, 2H, CH2), 3.70 (t, J = 5.9 Hz, 2H, CH2), 3.15 (s, 3H, CH3), 2.77 - 2.62 (m, 2H, CH2), 2.11 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.76, 161.88, 160.70, 160.23, 152.22, 150.55, 138.86, 133.21, 131.84, 131.72, 128.72, 127.58, 119.34, 117.70, 116.08, 97.77, 96.84, 57.38, 44.76, 43.93, 42.25, 28.45, 16.59. ESI-MS: m / z 518.39 (M+H) + ,C 27 H 27 N5O4S (517.18).

[0127] Example 34: (E)-3-(4-((2-(cyanomethyl)-l,2,3,4-tetrahydroisoquinolin-7-yl)amino)pyrimidin-4-yl)oxy)-3,5-dimethylphenyl)acrylonitrile (6e)

[0128]

[0129] Preparation method is same as example 30, only change the raw material to bromoacetonitrile. White solid, yield 81.2%. 1H NMR (600 MHz, DMSO-d6) δ 9.56 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.67 (d, J = 16.6 Hz, 1H, CH=), 7.55 (s, 2H, Ph-H), 7.15 (s, 1H, Ph-H), 7.07 (s, 1H, Ph-H), 6.83 (d, J = 8.3 Hz, 1H, Ph-H), 6.54 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 6.48 (d, J = 16.7 Hz, 1H, CH=), 3.86 (s, 2H, CH2), 3.24 (s, 2H, -CH2-CN), 2.71 (d, J = 5.5 Hz, 2H, -CH2), 2.68 (d, J = 5.4 Hz, 2H, -CH2), 2.11 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.63, 160.70, 160.25, 152.26, 150.29, 138.36, 138.35, 134.07, 131.77, 129.67, 128.80, 128.67, 126.22, 117.62, 116.34, 115.85, 97.59, 96.99, 54.80, 49.08, 43.36, 27.85, 17.00. ESI-MS: m / z 435.13 (M-H) - ,C 26 H 24 N6O (436.20).

[0130] Example 35: (E)-6-((4-(4-(2-cyanoethenyl)-2,6-dimethylphenoxy)pyrimidin-2-yl)amino)- 3,4-dihydroisoquinoline-2(lH)-carboxylic acid tert-butyl ester (7)

[0131]

[0132] Preparation method is same as example 2, only change the raw material to 6-amino-3,4- dihydroisoquinoline-2(lH)-carboxylic acid tert-butyl ester. Brown solid, yield 44.3%. 1H NMR (600 MHz, DMSO-d6) δ 9.58 (s, 1H, NH), 8.38 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.62 (d, J = 16.7 Hz, 1H, CH=), 7.51 (s, 2H, Ph-H), 7.21 (s, 2H, Ph-H), 6.85 (s, 1H, Ph-H), 6.54 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.41 (d, J = 16.4 Hz, 1H, CH=), 4.08 (s, 2H, CH2), 3.48 (t, J = 5.9 Hz, 2H, CH2), 2.61 (t, J = 5.9 Hz, 2H, CH2), 2.11 (s, 6H, CH3x2), 1.46 (s, 9H, CH3x3). ESI-MS: m / z 498.40 (M+H) + ,C 29 H 31 N5O3 (497.24).

[0133] Example 36: (E)-3-(3,5-dimethyl-4-(2-(1,2,3,4-tetrahydroisoquinolin-6-yl)amino)pyrimidin-4- yl)oxy)phenyl)acrylonitrile (8)

[0134]

[0135] Preparation method is same as example 29, only change the raw material to (E)-6-((4-(4-(2- cyanoethenyl)-2,6-dimethylphenoxy)pyrimidin-2-yl)amino)-3,4-dihydroisoquinoline-2(lH)- carboxylic acid tert-butyl ester 7. White solid, yield 82.3%. ESI-MS: m / z 398.23 (M+H) + ,C 24 H 23 N5O (397.19).

[0136] Example 37: (E)-3-(4-((2-(cyanomethyl)-l,2,3,4-tetrahydroisoquinolin-7-yl)amino)pyrimidin-4- yl)oxy)-3,5-dimethylphenyl)acrylonitrile (6e)

[0137]

[0138] Preparation method is same as example 30, only change the raw material to (E)-3-(3,5-dimethyl-4- (2-(l,2,3,4-tetrahydroisoquinolin-6-yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile 8. White solid, yield 82.3%. 1HNMR (400 MHz, DMSO-d6) δ 9.58 (s, 1H, NH), 8.39 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 7.66 (d, J = 16.7 Hz, 1H, CH=), 7.55 (s, 2H, Ph-H), 7.22 (s, 1H, Ph-H), 7.12 (s, 1H, Ph-H), 6.87 (s, 2H, NH2), 6.84 (s, 1H, Ph-H), 6.55 (d, J = 3.9 Hz, 1H, pyrimidyl-H), 6.48 (d, J = 18.4 Hz, 1H, CH=), 4.03 (s, 2H, piperidine-CH2), 3.19 - 3.11 (m, 2H, piperidine-CH2), 2.52 (s, 2H, piperidine-CH2), 2.10 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.76, 160.65, 160.15, 152.28, 150.51, 138.93, 133.63, 131.74, 129.75, 128.76, 126.75, 125.67, 119.34, 117.87, 117.28, 97.69, 96.70, 47.73, 43.15, 29.25, 14.59. ESI-MS: m / z 475.01 (M-H) - ,C 24 H 24 N6O3S (476.16).

[0139] Example 38: (E)-3-(4-((2-(cyanomethyl)-l,2,3,4-tetrahydroisoquinolin-7-yl)amino)pyrimidin-4-yl)oxy)-3,5-dimethylphenyl)acrylonitrile (6e)

[0140]

[0141] Preparation method is same as example 37, only change the raw material to methanesulfonyl chloride. White solid, yield 81.5%. 1HNMR (600 MHz, DMSO-d6) δ 9.59 (s, 1H, NH), 8.38 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.65 (d, J = 16.7 Hz, 1H, CH=), 7.53 (s, 2H, Ph-H), 7.22 (s, 1H, Ph-H), 7.11 (s, 1H, Ph-H), 6.87 (d, J = 8.5 Hz, 1H, Ph-H), 6.54 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.46 (d, J = 16.7 Hz, 1H, CH=), 4.20 (s, 2H, piperidine-CH2), 3.31 (d, J = 5.9 Hz, 2H, piperidine-CH2), 2.90 (s, 3H, CH3), 2.51 (s, 2H, piperidine-CH2), 2.09 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.75, 160.73, 160.21, 150.49, 139.08, 133.60, 131.80, 131.75, 128.74, 126.69, 125.32, 119.30, 118.10, 117.41, 97.75, 96.76, 46.07, 43.13, 33.87, 29.30, 14.60. ESI-MS: m / z 476.02 (M+H) + ,C 25 H 25 N5O3S (475.17).

[0142] Example 39: (E)-(6-((4-(4-(2-cyanoethenyl)-2,6-dimethylphenoxy)pyrimidin-2-yl)amino)-3,4-dihydroisoquinolin-2(lH- yl)dimethyl phosphonate (9c)

[0143]

[0144] Preparation method is same as example 37, only change raw material to dimethyl chlorophosphate. White solid, yield 75.2%. 1H NMR (600 MHz, DMSO-d6) δ 9.54 (s, 1H, NH), 8.37 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.65 (d, J = 16.7 Hz, 1H, CH=), 7.53 (s, 2H, Ph-H), 7.17 (s, 1H, Ph-H), 7.04 (s, 1H, Ph-H), 6.81 (d, J = 8.3 Hz, 1H, Ph-H), 6.53 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.46 (d, J = 16.7 Hz, 1H, CH=), 4.07 (d, J = 6.3 Hz, 2H, piperidine-CH2), 3.55 (s, 3H, -OCH3), 3.53 (s, 3H, -OCH3), 3.20 (d, J = 9.0 Hz, 2H, piperidine-CH2), 2.35 (s, 2H, piperidine-CH2), 2.09 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.75, 160.73, 160.22, 152.30, 150.50, 138.69, 134.26, 131.78, 131.71, 128.74, 126.88, 126.23, 119.30, 118.52, 117.23, 97.61, 96.69, 52.97, 45.70, 42.10, 29.38, 16.55. ESI-MS: m / z 506.02 (M+H) + ,C 26 H 28 N5O4P (505.19).

[0145] Example 40: (E)-3-(3,5-dimethyl-4-(2-(2-(methylsulfonyl)acetyl)-l,2,3,4- tetrahydroisoquinolin-6-yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (9d)

[0146]

[0147] Preparation method is same as example 37, only change the raw material to 2- methylsulfonylacetyl chloride. White solid, yield 81.6%. 1H NMR (400 MHz, Chloroform-d) δ 9.59 (s, 1H, NH), 8.38 (d, J = 5.7 Hz, 1H, pyrimidyl-H), 7.65 (d, J = 16.8 Hz, 1H, CH=), 7.53 (d, J = 10.3 Hz, 2H, Ph-H), 7.22 (s, 1H, Ph-H), 7.12 (s, 1H, Ph-H), 6.87 (d, J = 8.3 Hz, 1H, Ph-H), 6.53 (d, J = 5.5 Hz, 1H, pyrimidyl-H), 6.46 (d, J = 16.7 Hz, 1H, CH=), 4.59 (s, 1H, CH2x ½), 4.51 (s, 3H, CH2; CH2x ½), 3.62 (m, 2H, CH2), 3.12 (s, 3H, CH3), 2.58 (s, 1H, CH2x ½), 2.37 (s, 1H, CH2x ½), 2.09 (s, 6H, CH3x 2). 13 C NMR (100 MHz, DMSO-d6) δ 168.74, 161.82, 161.71, 160.72, 160.22, 152.25, 150.48, 139.03, 134.74, 131.79, 128.71, 126.67, 126.51, 119.36, 118.01, 117.40, 97.74, 96.85, 57.44, 47.26, 42.25, 30.26, 28.87, 16.00. ESI-MS: m / z 518.47 (M+H) + ,C 27 H 27 N5O4S (517.18).

[0148] Example 41: (E)-3-(3,5-dimethyl-4-(2-(2-(methylsulfonyl)ethyl)-1,2,3,4-tetrahydroisoquinolin-6-yl)amino)pyrimidin-4-yl)oxy)phenyl)acrylonitrile (9e)

[0149]

[0150] Preparation method is same as example 37, only change raw material to dimethyl chlorophosphate. White solid, yield 75.2%. 1H NMR (600 MHz, DMSO-d6) δ 9.51 (s, 1H, NH), 8.36 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 7.64 (d, J = 16.7 Hz, 1H, CH=), 7.52 (s, 2H, Ph-H), 7.14 (s, 1H, Ph-H), 7.03 (s, 1H, Ph-H), 6.73 (d, J = 8.3 Hz, 1H, Ph-H), 6.51 (d, J = 5.6 Hz, 1H, pyrimidyl-H), 6.45 (d, J = 16.7 Hz, 1H, CH=), 3.47 (s, 2H, piperidine-CH2), 3.35 (t, J = 6.8 Hz, 2H, CH2), 3.00 (s, 3H, CH3), 2.84 (t, J = 6.8 Hz, 2H, CH2), 2.60 (t, J = 5.9 Hz, 2H, piperidine-CH2), 2.43 (s, 2H, piperidine-CH2), 2.09 (s, 6H, CH3x2). 13 C NMR (100 MHz, DMSO-d6) δ 168.74, 160.68, 160.26, 152.30, 150.54, 138.55, 134.22, 131.76, 131.73, 128.72, 127.89, 126.55, 119.38, 118.07, 116.90, 97.51, 96.71, 55.03, 51.73, 51.40, 50.63, 42.03, 29.67, 16.56. ESI-MS: m / z 504.00 (M+H) + ,C 27 H 29 N5O3S (503.20).

[0151] Example 42: Anti-HIV activity test (MT-4 cell model)

[0152] See ① Pauwels R, et al. J. Virol. Methods. 1988, 20, 309. ② Pannecouque C, et al. Nat Protocols 2008, 3, 427.

[0153] Terminology

[0154] MT-4 cells: human acute lymphoblastic leukemia cells; MTT assay: MTT is 3-(4,5-dimethylthiazol-2)-2,5-diphenyltetrazolium bromide; trade name: thiazolyl blue; DMSO: dimethyl sulfoxide.

[0155] Test principle

[0156] Since HIV-infected MT-4 cells will be cytopathic in a certain period of time (5-7 days), a proper concentration of the solution of the compound to be tested is added to the suspension of HIV-infected MT-4 cells, and after a period of time (5-7 days) of culture, the cell viability is determined by MTT assay, and the concentration of the drug that protects 50% of the cells from cytopathic effect (EC 50 ) is obtained, and the anti-HIV activity of the target compound is determined. At the same time, the concentration of the target compound that causes 50% of the uninfected HIV cells to be cytopathic (CC 50 ) is obtained. Principle of MTT assay: MTT, i.e. bromide-3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium nitrogen, can be combined with succinic acid dehydrogenase in living cells, but not with dead cells. MTT assay is a rapid enzyme assay that reflects cell viability.

[0157] Test materials and methods

[0158] (1) HIV-1 wild strain III B , single mutant strains L100I, K103N, Y181C, Y188L, E138K, and double mutant strains K103N / Y181C (RES056) and F227L / V106A, HIV-2 ROD strain: provided by the Institute of Microbiology and Immunology, Rega Institute, Leuven University, Belgium.

[0159] (2) MT-4 cells: provided by the Institute of Microbiology and Immunology, Rega Institute, Leuven University, Belgium.

[0160] (3) MTT: purchased from Sigma Company, USA.

[0161] (4) Sample treatment: the sample is dissolved in DMSO to prepare a proper concentration before use, and double distilled water is used for 5-fold dilution, and each dilution is 5 times.

[0162] (5) Positive control drugs: doravirine (DOR), nevirapine (NVP), efavirenz (EFV), and etravirine (ETV).

[0163] (6) Test method: after dilution of the sample, it is added to the suspension of HIV-infected MT-4 cells, and after a period of time, the cell viability is determined by MTT colorimetric method, and the absorbance (A) value is recorded at 590 nm in the enzyme marker instrument, and the EC 50 , CC 50 and SI are calculated.

[0164] (7) MTT staining: After incubation with the test compound for a certain period of time, 20 μL of MTT solution was added to each well, and the incubation was continued for several hours. The staining solution was then discarded, and 150 μL of DMSO was added to each well. The absorbance was then measured at 590 nm using a microplate reader.

[0165] The procedure was as follows: the compound was dissolved in DMSO or water and diluted with phosphate buffer. 3 x 10 5 MT-4 cells were preincubated with 100 μL of different concentrations of the test compound for 1 h at 37°C. Then, 100 μL of an appropriate dilution of the virus was added to the mixture, and the cells were incubated for 1 h at 37°C. After three washes, the cells were resuspended in culture medium with or without the test compound. The cells were then incubated for 7 days at 37°C in a 5% CO2atmosphere, and the original culture medium was replenished with or without the test compound on the third day after infection. Each culture condition was repeated twice. The cytopathic effect of the virus was monitored daily using an inverted light microscope. In general, the virus dilution used in this experiment caused cytopathic effect in the cells on the fifth day after infection. The inhibitory concentration of the drug was expressed as the concentration at which the drug produced 50% inhibition of the cytopathic effect of the virus while being non-toxic to the cells (EC 50 ). It is important to emphasize that when the water solubility of the compound was poor, DMSO was used to dissolve the compound. In this case, the volume ratio of DMSO to water was generally less than 10% (the final concentration of DMSO in the MT-4 cell culture medium was less than 2%). Because DMSO can affect the antiviral activity of the test compound, the antiviral activity of the test compound in the presence of DMSO was compared with the antiviral activity of the test compound in the absence of DMSO. In addition, the final concentration of DMSO (1 / 1000) was much lower than the concentration required to affect the replication of HIV-1 in MT-4 cells.

[0166] The in vitro anti-HIV activity screening data of the target compounds were provided by the Institute of Microbiology and Immunology, Rega Institute for Medical Research, University of Leuven, Belgium. All the activity data were measured in at least three independent and parallel experiments.

[0167] First, we tested the antiviral activity and cytotoxicity of all the compounds against HIV-1 wild strain and RES056 mutant strain, and HIV-2 ROD strain. As shown in Table 1, the diarylpyrimidine compounds provided by the present application containing a fused heterocycle exhibited good anti-HIV-1 wild strain activity, with an EC 50 range of 0.0018 to 0.12 μM. Among them, compound 3a (EC 50 = 0.0019 μM), 3d (EC 50 = 0.0019 μM), and 3k (EC 50=0.0019μM), 3s(EC) 50 =0.0020μM) and 3u(EC) 50 The activity of the drug at 0.0018 μM was significantly superior to all control drugs (EC). 50 =0.0032–0.13 μM); for the HIV-1RES056 mutant strain, 3k (EC) 50 =0.35μM) showed the highest antiviral activity, superior to the marketed drug NVP (EC). 50 >9.51μM), comparable to EFV (EC). 50 =0.36 μM); for HIV-2 ROD strain, the three compounds 3a, 3k, and 3r exhibited micromolar-level antiviral activity (EC). 50 =1.26–7.04 μM), significantly superior to all control drugs (which showed no antiviral activity at the tested concentrations).

[0168] Table 1. Activity and cytotoxicity of compounds against wild-type and RES056 mutant HIV-1 and HIV-2.

[0169]

[0170]

[0171] Note: a EC 50 The concentration of compounds that protect 50% of HIV-infected MT-4 cells from cytopathic effects; b CC 50 The concentration at which the target compound causes 50% of uninfected HIV cells to become diseased; c SI: Selection Index, CC 50 With EC 50 The ratio; d NA: No result can be calculated.

[0172] Compounds exhibiting excellent antiviral activity in the initial testing were further tested for their activity against HIV-1 mutant strains L100I, K103N, Y181C, Y188L, E138K, and F227L / V106A. As shown in Table 2, compound 3k(EC... 50 =0.0019 μM) showed particularly outstanding activity against the K103N mutant strain, superior to all positive control drugs (EC). 50= 0.0030 - 5.22 μM); for mutant strains L100I, Y181C, E138K, the activities were 0.0087, 0.032 and 0.011 μM, respectively, which were superior to or equivalent to the positive control drug. Therefore, the compounds of this type have great value for research and development, and can be used as candidate drugs for the preparation of anti-HIV.

[0173] Table 2 Antiviral activity of compounds against HIV-1 clinical common mutant strains

[0174]

[0175] Note: a EC 50 : Compound concentration for protecting 50% of HIV-1 infected MT-4 cells from cytopathic effect.

[0176] Example 43: Anti-CHIKV activity test (SJCRH30 cell model)

[0177] See ① Gupta DK, et al. Mar Drugs. 2014, 12(1), 115-27.

[0178] Test principle

[0179] Alamar-Blue Cell Viability Assay Kit is a kit for detecting cell proliferation by redox indicator Resazurin. Resazurin is in the oxidized state, showing blue and basically no fluorescence. When it is taken in by cells or bacteria and metabolized by enzymes, it will be converted into pink reduction product Resorufin. Resorufin can emit strong red fluorescence, and the change in cell viability can be detected by fluorescence.

[0180] Plaque assay is an experimental method for determining viral infectivity. Appropriate concentration of virus suspension is added to the layered monolayer cell culture to allow virus adsorption, and then a layer of melted agar is covered. Each virus replicates in infected cells to produce a localized focus of infection, i.e. plaque. Live cells can be seen as colorless plaques by staining with neutral red. Each plaque is produced by the replication of an infectious virion, called plaque forming unit (PFU), i.e. one plaque is equivalent to one virion. The amount of infectious virus contained in the virus suspension is expressed in terms of plaque forming units per milliliter, i.e. PFU / ml.

[0181] Test materials and methods

[0182] (1) Cell and compound preparation: BHK21 (juvenile hamster kidney) cells and SJCRH30 rhabdomyosarcoma cells (ATCCCRL-2061) were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (FCS) at 37°C and 5% CO2. C6 / 36 mosquito cells isolated from Aedes albopictus embryonic tissue were cultured in L-15 medium containing 10% FCS at 28°C. CHIKV-122508 (SGEHICHD 122508, provided by the Singapore Institute of Environmental Health) was multiplied in C6 / 36 cells and stored at -80°C before use. Compounds were dissolved in DMSO to a final stock concentration of 10 mM and stored at -80°C.

[0183] (2) Cell viability assay: SJCRH30 cells were seeded in 96-well cell culture plates and incubated overnight at 37°C and 5% CO2. The compound was diluted with 2% FCS and then incubated with a monolayer of SJCRH30 cells at 37°C and 5% CO2 for 24 hours. After the incubation period, the cells were washed once with phosphate-buffered saline (PBS), and 10 μL was added to each well. The reagents were then incubated at 37°C and 5% CO2 for 4 hours. A microplate reader (Tecan) was used. The fluorescence detected was read at an excitation wavelength of 570 nm and an emission wavelength of 600 nm (Switzerland). Measurements of cells treated with the compound or 0.1% DMSO solvent (control) were normalized to measurements of untreated cells. Cytotoxic concentration (CC) 50 () is defined as the treatment concentration at which a compound causes 50% of cells to become diseased.

[0184] (3) Drug inhibition study: SJCRH30 cells were seeded into 96-well plates and incubated overnight at 37°C and 5% CO2. Cells were then infected with 1 MOI (multiple of infection) of CHIKV-122508 for 1.5 hours. Subsequently, the cells were washed twice with PBS, and the desired drug concentration was added to the cells and incubated for 24 hours. The supernatant from the CHIKV-infected cells was then extracted for viral plaque assays. Three independent experiments were performed for each drug concentration group.

[0185] (4) Viral Plaque Assay: BHK cells were seeded in 24-well plates and incubated overnight at 37°C and 5% CO2. The CHIKV-treated supernatant was serially diluted 10-fold from 10¹ to 10⁶, and 100 μL of the diluted supernatant was added to the BHK cells. Incubation was performed for 1.5 hours, with shaking at 15-minute intervals during this period. After incubation, the plates were washed twice with PBS and covered with 1% carboxymethyl cellulose (CMC). The assay plates were incubated at 37°C and 5% CO2 for 3 days. The CMC was removed, and the cells were stained and fixed with 10% paraformaldehyde-1% crystal violet (Sigma-Aldrich Chemical, St. Louis, MO, USA) solution for observation and plaque counting. Viral titers were expressed as plaque-forming units (PFU) per milliliter.

[0186] The in vitro anti-CHIKV activity screening data of the target compounds were provided by the Yong Loo Lin School of Medicine, National University of Singapore. All activity data were obtained through at least three independent, parallel experiments.

[0187] First, the cytotoxicity of the compounds was tested using an Almar Blue cell viability assay kit. Compounds with cell viability exceeding 80% at a 10 μM concentration were further tested for their anti-CHIKV activity. As shown in Table 3, compounds 3b, 3g, 3j, 3k, 3q, 3r, 6a, and 9d exhibited good anti-CHIKV activity at a 10 μM concentration. In particular, compounds 3q and 3r showed inhibitory activity of 3.4-log and 3.1-log, respectively, compared to the negative control (0.1% DMSO). This is the first report of anti-CHIKV activity in diarylpyrimidine derivatives containing fused heterocycles. Given that there are currently no approved drugs specifically targeting CHIKV, these compounds have significant research and development value and can be utilized as candidate drugs for the preparation of anti-CHIKV drugs.

[0188] Table 3. Activity of compounds in inhibiting CHIKV replication

[0189] Compounds a Inhibition at 10 μM ​ 3b 1.61-log 3g 1.98-log 3j 1.99-log 3k 1.41-log 3q 3.4-log 3r 3.1-log 6a 1.74-log 9d 1.86-log

[0190] a Compared to 0.1% DMSO, the compound exhibited logarithmic inhibition of CHIKV at a concentration of 10 μM.

[0191] IV. Conclusion

[0192] The di-aryl pyrimidine derivative containing fused heterocycle provided by the present application is a series of novel compounds. As shown in Table 1, the activities of five compounds 3a, 3d, 3k, 3s and 3u against HIV-1 wild strain are obviously superior to all the control drugs; for HIV-2 ROD strain, the antiviral activities of compounds 3a, 3k and 3r are significantly superior to all the control drugs. As shown in Table 2, the activity of compound 3k against K103N mutant strain is significantly superior to all the positive control drugs. As shown in Table 3, compared with the negative control (0.1% DMSO), compounds 3q and 3r exhibit significant anti-CHIKV activity at a concentration of 10 μM. Therefore, the compounds have great value for research and development, and can be utilized and developed as candidate drugs for preparing anti-HIV and anti-CHIKV.

Claims

1. A diarylpyrimidine compound containing a fused heterocycle or a pharmaceutically acceptable salt thereof, having a structure as shown in the following general formula I: wherein, R1 is CN, CH=CHCN; R2 is H, NH2, F, Br, I, Cl; R3 is H, NH2, F, Br, I, Cl; X is The diarylpyrimidine compound containing a fused heterocycle is one of the following: The pharmaceutically acceptable salt of the compound is a hydrochloride, a sulfate, a tartrate or a citrate. The preparation method of the diarylpyrimidine derivative containing a fused heterocycle comprises the following steps: first, using 2,4-dichloropyrimidine 1 as a raw material, an intermediate 2 is generated by reacting with (E)-3,5-dimethyl-4-hydroxyphenylacrylonitrile in an N,N-dimethylformamide solution; the intermediate 2 is coupled with an aniline of a fused heterocycle under the catalysis of palladium acetate to obtain a target compound 3(a-x) or an intermediate 4 or 7; then, the intermediate 4 or 7 is removed from a Boc group to obtain an intermediate 5 or 8, and the intermediate 5 or 8 is reacted with different sulfonyl chlorides, phosphoryl chlorides, acyl chlorides or halogenated alkanes to obtain final products 6(a-e) or 9(a-e); Reagents and conditions: (i) (E)-3,5-dimethyl-4-hydroxyphenylacrylonitrile, N,N-dimethylformamide, potassium carbonate, 50 °C; (ii) palladium acetate, 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene, cesium carbonate, nitrogen, 1,4-dioxane, 90 °C; (iii) trifluoroacetic acid, dichloromethane, room temperature; (iv) sulfonyl chloride, phosphoryl chloride, acyl chloride or halogenated alkane, triethylamine, 0 °C.

5. Use of the diarylpyrimidine compound containing a fused heterocycle according to claim 1 in the preparation of an anti-HIV drug.

2. The fused heterocyclic ring-containing diarylpyrimidines of claim 1, wherein 6. Use of the diarylpyrimidine compound containing a fused heterocycle according to claim 2 in the preparation of an anti-Chikungunya virus drug; the diarylpyrimidine compound containing a fused heterocycle is selected from compounds 3a, 3b, 3c, 3g, 3q, 3k, 3r, 6a, 6b, 9a, 9b.

3. The fused heterocyclic ring-containing diarylpyrimidines of claim 1, wherein 7. A pharmaceutical composition comprising the diarylpyrimidine compound containing a fused heterocycle according to claim 1 and one or more pharmaceutically acceptable carriers.

4. The process for preparing a fused heterocycle-containing diarylpyrimidine compound according to claim 2, wherein the step of ​ ​ ​ ​ ​ ​

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