CN114539163B - HIV-1 reverse transcriptase inhibitor and its synthesis process - Google Patents

HIV-1 reverse transcriptase inhibitor and its synthesis process Download PDF

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CN114539163B
CN114539163B CN202210147690.8A CN202210147690A CN114539163B CN 114539163 B CN114539163 B CN 114539163B CN 202210147690 A CN202210147690 A CN 202210147690A CN 114539163 B CN114539163 B CN 114539163B
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孟歌
张凤
程亚楠
陈芬儿
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Abstract

The invention belongs to the technical field of medicines, and in particular relates to an HIV-1 reverse transcriptase inhibitor and a synthesis method thereof. The HIV-1 reverse transcriptase inhibitor is a substituted aryl urea imino diaryl pyrimidine derivative, and also comprises medicinal salts, hydrates and solvates thereof, polycrystal or eutectic crystal thereof, and precursors and derivatives with the same biological functions. The results of in vitro cell level anti-HIV-1 activity experiments show that the compounds have stronger anti-HIV-1 biological activity, can obviously inhibit MT-4 intracellular virus replication infected by HIV-1 virus, and have lower cytotoxicity and better selectivity. The invention also comprises the application of the composition of the compound in medicaments related to the treatment of AIDS and the like.

Description

HIV-1 reverse transcriptase inhibitor and its synthesis process
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to an HIV-1 reverse transcriptase inhibitor and a synthesis method thereof.
Background
The reverse transcription process (Reverse transcription) is the primary link after the invasion of the host cell by the HIV-1 virus and plays a key role in the HIV-1 life cycle. In this link, reverse transcriptase (Reverse transcriptase, RT) is responsible for reverse transcription of viral RNA into DNA-RNA hybrids and RNA degradation in the hybrids to form single stranded viral DNA, which is then integrated into host cells by integrase. Thus, RT can be used as an important selective target for drug design against HIV-1 virus 1 。
Among the existing targeted HIV-1 reverse transcriptase compounds, non-nucleoside reverse transcriptase inhibitors (NNRTIs) have stronger antiviral activity and lower cytotoxicity 2 Thus becoming the first drug for high-efficiency antiretroviral therapy (HAART) cocktail therapy of anti-HIV-1 treatment 3 . Five anti-HIV reverse transcriptase inhibitors are approved by the FDA in the united states: nevirapine (Nevirapine), delavirdine (Delavirdine), efavirenz (Efavirenz), itravirenz (Etravirine), rilpivirine (Rilpivirine). In addition, RDEA806, IDX899, UK-453061 are undergoing clinical studies 4 . However, amino acid mutations in reverse transcriptaseCan deactivate the originally effective drug, namely generate drug-resistant HIV virus strain 5 . Therefore, the development of novel high-efficiency non-nucleoside reverse transcriptase inhibitors with broad-spectrum drug resistance is one of the hot spots of research by pharmaceutical chemists 6 。
The invention aims at carrying out structural optimization on itravirin and rilpivirine, and designs and synthesizes a series of diaryl pyrimidine derivatives containing substituted aryl urea imino groups by carrying out structural modification on linking groups of a left wing aromatic ring and a middle pyrimidine ring of the compounds, and introduces various substituents on the aromatic ring, so as to enhance interaction between the compounds and conserved aromatic amino acids on the inner wall of a binding pocket of a reverse transcriptase non-nucleoside drug, and further improve the biological activity of the series of compounds against drug-resistant virus strains.
Disclosure of Invention
The invention aims to provide an HIV-1 non-nucleoside reverse transcriptase inhibitor with excellent performance and a synthesis method thereof.
The HIV-1 reverse transcriptase inhibitor provided by the invention is a diaryl pyrimidine derivative containing substituted aryl urea imino, and has a structural formula shown in the following formula (I):
Figure BDA0003509581640000011
wherein R is 1 Independently selected from hydrogen, methyl, cyano, nitro, methoxy, ethoxy, hydroxy, halogen, the substitution position may be ortho, para or meta; x is halogen.
The HIV-1 reverse transcriptase inhibitor also comprises medicinal salts, hydrates and solvates of the diaryl pyrimidine derivatives containing substituted aryl urea imino, polycrystal and eutectic of the diaryl pyrimidine derivatives, and precursors and derivatives with the same biological functions.
The pharmaceutically acceptable salts include in particular hydrochloride, hydrobromide, sulfate, phosphate, acetate, mesylate, p-toluenesulfonate, tartrate, citrate, fumarate or malate salts.
The invention also provides a synthetic method of the HIV-1 reverse transcriptase inhibitor-substituted aryl ureido diaryl pyrimidine derivative, which comprises the following synthetic route:
Figure BDA0003509581640000021
the specific steps of the synthesis are as follows;
firstly, using cheap and easily available thiouracil (5, 1.5-1.6 equiv.) as a starting material, using methyl iodide (6,1.0-1.1 equiv.) as a methylation reagent, reacting for 20-24 h under the action of sodium hydroxide (about 1.25M and 1.0-1.1 equiv.) at room temperature, and carrying out S-alkylation reaction to obtain a high-purity white solid, namely 2-methylthiopyrimidine-4-ketone (7), wherein the yield is 89.7% 7 ；
(II) 2-methylthio pyrimidine-4-ketone (7,1.0-1.1 equiv.) and excessive 4-cyanoaniline (8,2.5-3.0 equiv.) are reacted for 10-18 h in a molten state at 180-185 ℃ under the condition of no solvent, and after acetonitrile dissolution, appropriate post treatment is carried out, thus obtaining yellow solid 2- (4-cyanoanilino) pyrimidine-4-ketone (9) with the yield of 67.6 percent 8 ；
Dissolving and refluxing 2- (4-cyanoanilino) pyrimidine-4-ketone (9,1.0-1.1 equiv.) in a large excess of phosphorus oxychloride (10.0-11.0 equiv.), performing chlorination reaction on hydroxyl at C-4 position of pyrimidine heterocycle, performing post-treatment, dissolving in a proper amount of cold water, neutralizing to be neutral by sodium hydroxide (20%), obtaining yellow precipitate, filtering and drying to obtain yellow solid 2- (4-cyanoanilino) -4-chloro-pyrimidine (10), wherein the yield is 84.0%, and the compound is an important intermediate for synthesizing a target compound;
(IV) nucleophilic substitution reaction of 2- (4-cyanoanilino) -4-chloro-pyrimidine (10,1.0-1.1 equiv.) with 2-halogenobenzonitrile (11 a,11b, 1.5-1.6 equiv.) under the action of sodium hydride (60%, 2.0-2.2 equiv.) in dried N, N-dimethylformamide (2.5-3.0 equiv. ML), anhydrous and anaerobic conditions to obtain unstable intermediate Cyan-CH 2 -DAPYs(12a,12b)。；
(V) because of the above intermediate Cyan-CH 2 DAPYs (12 a,12 b) are notStable, can remove nitrogen protection after reaction, and can be placed in air to react for 48-72h under room temperature condition, can slowly oxidize to obtain key intermediate Oxo-CH 2 DAPYs, and obtaining 2-halogenophenyl 2- (4-cyanophenylamino) -pyrimidinone pure products (13 a,13 b) through post-treatment and column chromatography separation;
and (six) simultaneously carrying out the steps (one) - (five), taking various substituted anilines as starting materials, and carrying out the preparation of various substituted semicarbazide (18 a-18 t) in parallel through two steps of reactions. The specific method is that various substituted anilines (14 a-14t, 1.0-1.1 equiv.) are dissolved in tetrahydrofuran, and NaHCO is simultaneously dissolved 3 (1.2-1.4 equiv.) dissolving in water, and mixing NaHCO with water 3 The aqueous solution was mixed with a tetrahydrofuran solution of a substituted aniline and placed in an ice bath, and phenyl chloroformate (15,1.2 to 1.4 equiv.) was added after the temperature had stabilized at 0 to 5 ℃. The reaction speed is extremely high, and the reaction can be completed after the reactants are added. Because the substituted phenyl carbamate intermediate is unstable in the reaction liquid, the temperature in the reaction process is controlled to be 0-5 ℃ so as to prevent the intermediate from being decomposed and affecting the yield and purity. The post-treatment process is simple, and various stable stituted phenyl carbamate intermediates at room temperature can be obtained after extraction and rotary evaporation by ethyl acetate 9 . The substituted phenyl carbamate intermediate (16 a-16t, 1.0-1.1 equiv.) is dissolved in acetonitrile, 80% hydrazine hydrate (17,2.5-3.0 equiv.) is added, and the reaction is carried out at room temperature under ultrasound for 1-3 h to obtain the corresponding substituted semicarbazide (18 a-18 t). The method comprises the steps of carrying out a first treatment on the surface of the
(seventh) finally, the intermediate Oxo-CH 2 DAPYs (13 a,13b, 1.0-1.1 equiv.) and various substituted semicarbazide (18 a-18t, 1.0-1.1 equiv.) are heated, refluxed and dehydrated in ethanol for 4-5 h under the condition of taking hydrochloric acid as a catalyst, and then the corresponding target compound (1 a,1b, …,1z,1 aa) is obtained.
In step six, the substituted anilines (14 a-14 t), substituted phenyl carbamates intermediates (16 a-16 t), substituted semicarbazides (18 a-18 t), and 27 targets (1 a,1b, …,1z,1 aa) obtained in step seven, corresponding X, R, are listed below:
Figure BDA0003509581640000031
Figure BDA0003509581640000041
the numbers (a-v) corresponding to substituents in the table correspond to the respective series of intermediates (14, 16, 18) and the respective series of target compounds (1 a-v), respectively, and the intermediate numbers and substituent type numbers are combined with each other to represent the respective numbers of the respective compounds, such as the intermediates (14 a-v, 16 a-v, 18 a-v) and the target compounds (1 a-v), respectively.
The compound (I) is an HIV-1 non-nucleoside reverse transcriptase inhibitor, has stronger biological activity, higher selection coefficient and smaller cytotoxicity.
The invention also relates to a pharmaceutical composition containing an effective dose of the compound (I) and a related medicinal carrier, and application of the compound or the composition in preparing medicines for preventing and treating AIDS.
In summary, according to the structural characteristics of the listed medicines, namely itravirin and Li Piwei, and according to the structure-activity relationship of CH-DAPYs, the main pharmacophore is reserved, and the structure of semicarbazone with antiviral activity is introduced onto the methylene connecting group of the left wing aromatic ring of the DAPY compound by utilizing the split principle, so as to design and synthesize a series of diaryl pyrimidine compounds with semicarbazone structures. Molecular modeling shows that: the compound is in a classical U-shaped conformation when combined with reverse transcriptase, a left wing aromatic ring enters a hydrophobic region, pi-pi stacking effect is formed with Y181 and Y188, and NH connecting a right wing aromatic ring and a pyrimidine ring can form a hydrogen bond with K101. Attempts have been made to thereby increase the interaction of the compound with the target in an attempt to increase the biological activity of the compound of interest against drug-resistant HIV strains. The invention uses thiouracil and methyl iodide as initial raw materials, and the target compound is obtained through S-methylation, nucleophilic substitution, chlorination, oxidation, second nucleophilic substitution and condensation reaction in sequence. All new compounds are passed through 1 H NMR 13 C NMR and other spectroscopic analysis methods, and some compounds were confirmed by ESI-MS. In vitro cell of the target CompoundScreening experiments for anti-HIV-1 activity at both level and enzyme level. The cell level activity test result shows that the target compound has low micromolar inhibition activity on HIV-1 wild type strains, and part of the compounds have low inhibition activity on double mutant HIV-1 strains RES056 and HIV-2 ROD. Wherein the target compound has optimal 1h activity (EC 50 A value of 0.0329. Mu.M, an SI value of 3712), the activity is stronger than that of the reference drugs NEV and DEV, and the cytotoxicity is small and the selectivity is high. The test result of enzyme level on HIV-1RT activity shows that the target compound has strong inhibition activity on HIV-1RT.
Drawings
Figure 1 shows the strategy for designing the target compounds of the present invention from the marketed drugs itravirin and Li Piwei.
FIG. 2 shows the binding pattern of the target to the HIV-1RT non-nucleoside inhibitor binding pocket. Wherein, (a) target 1q, (b) target 1p, (c) target 1k, and (d) target 1d.
Detailed Description
The present invention will be better understood by the following examples of embodiments, but is not limited thereto.
Example 1: preparation of intermediates
(1) Preparation of 2- (4-cyanoanilino) -4-chlorouracil (10)
Sodium hydroxide (50.4 g,1.26 mol) was added to an aqueous solution (1.0L), and 2-thiouracil (5, 212.9g,1.50 mol) was added to the sodium hydroxide solution, stirred until completely dissolved, allowed to stand, and cooled to room temperature. Methyl iodide (6,153.6 g,1.2 mol) was added and stirring was continued for 24h, and thin layer chromatography showed complete conversion of the starting material. The pH is adjusted to be neutral, white solid is separated out, filtered, washed and dried to obtain white solid (153 g, 89.7%), namely 2-methylthiopyrimidine-4-ketone (7).
2-methylthiopyrimidin-4-one (7, 42.7g,0.30 mol) and p-aminobenzonitrile (8, 88.6g,0.75 mol) were mixed thoroughly, and the temperature was slowly raised to 180℃for reaction for 10 hours. Thin layer chromatography showed that a small amount of raw materials were not converted at 10h, the reaction time was prolonged to 18h, the raw materials were completely converted, but the product became heterogeneous, and the yield was rather lowered. After the reaction was completed, the reaction mixture was slowly cooled to room temperature, and the reaction mixture was solidified to form a pale yellow solid, which was stuck to the bottom of the flask, and was slurried with acetonitrile to give a crude 2- (4-cyanoanilino) pyrimidin-4-one (9, 43.0g, 67.6%).
The 2- (4-cyanoanilino) pyrimidin-4-one (9, 32.0g,0.165 mol) obtained above was dissolved in phosphorus oxychloride (150 mL,1.64 mol), which was now used as both a chloro reagent and a solvent. Heating and refluxing for 30min, and completely reacting. Cooling to room temperature, pouring into ice water slowly under intense stirring, controlling temperature, and otherwise, easily initiating detonation and boiling to affect the purity of the product. A pale yellow solid precipitated, cooled, filtered, and the solid was suspended in (200 mL) cold water and neutralized to neutrality with 20% sodium hydroxide. Filtration, washing with water and drying gave 2- (4-cyanoanilino) -4-chlorouracil (10, 32.0g, 84.0%) as a pale yellow solid.
(2) Preparation of 2-halophenyl 2- (4-cyanophenylamino) -pyrimidinone (13 a,13 b)
2- (4-cyanoanilino) -4-chlorouracil (10, 2.77g,12.0 mmol) was aromatically nucleophilic substituted with o-halophenylacetonitrile (11 a,11b,2.73g/3.53g,18.0 mmol) in the presence of NaH (0.96 g,24.0mmol, 60%) in anhydrous DMF (30 mL) to give 2- (p-cyanoanilino) -4- (2-halophenylcyano) methylenepyrimidine intermediate (12 a,12 b). The reaction is sensitive to air, requires nitrogen protection, and requires high anhydrous conditions, and DMF is used after drying overnight with activated molecular sieves. 2- (p-cyano-phenylamino) -4- (2-chlorphenyl cyano) methylene pyrimidine (12 a,12 b) is unstable, nitrogen protection is removed, and 2- (p-cyano-phenylamino) -4-aroyl pyrimidine (13 a,13 b) is obtained by oxidation in air. The reaction is a speed-limiting reaction in the whole synthesis, the time consumption is long, the side reaction is more, and the reaction is not easy to control. After 48-72h of reaction, TLC showed complete reaction. The reaction solution was poured into water, neutralized to neutrality with dilute hydrochloric acid, extracted with ethyl acetate, and the ethyl acetate layers were combined, dried over anhydrous sodium sulfate and removed by rotary evaporation, and the crude product obtained was subjected to flash column chromatography (PE: ea=5:1) to give pure 2-halophenyl 2- (4-cyanophenylamino) -pyrimidinone (13 a,13 b).
(3) Synthesis of various substituted semicarbazides
The substituted semicarbazide is obtained by taking various substituted anilines as starting materials and two steps. The method has the advantages of simple and feasible route, mild condition, economy, high efficiency, high yield and high purity.
Various substituted anilines (14 a-t, 1.86-3.44 g,20.0 mmol) were dissolved in tetrahydrofuran while NaHCO was added 3 (2.0 g,24.0 mmol) was dissolved in water and NaHCO was added 3 The aqueous solution was mixed with a tetrahydrofuran solution of the substituted aniline and placed in an ice bath, and phenyl chloroformate (15, 3.8g,24.0 mmol) was added after the temperature had stabilized at 0 ℃. The reaction speed is extremely high, and the reaction can be completed after the reactants are added. Because the substituted phenyl carbamate intermediate is unstable in the reaction liquid, the temperature in the reaction process is controlled at 0 ℃ so as to prevent the intermediate from being decomposed and influencing the yield and purity. The post-treatment process is simple, ethyl acetate is used for extraction and then rotary evaporation is carried out to obtain various substituted phenyl carbamate intermediates, and the intermediates are stable at room temperature 9 。
The substituted phenyl carbamate intermediate (16 a-t, 2.13-2.92 g,10.0 mmol) was dissolved in acetonitrile, 80% hydrazine hydrate (17, 1.6g,25.0 mmol) was added and the reaction was carried out at room temperature with ultrasound for 1-3 h to give the corresponding substituted semicarbazide (18 a-t), the specific experimental numbers are shown in Table 1.
TABLE 1 physical Properties and yields of various intermediate substituted semicarbazides 18a-t
Figure BDA0003509581640000061
Figure BDA0003509581640000071
Example 2: DAPY target 1a-aa synthesis
Substituted semicarbazide (18 a-t, 0.23-0.35 g,1.5 mmol) and 2-chlorophenyl 2- (4-cyanophenylamino) -pyrimidinone (13 a,13b,0.50g/0.57g,1.5 mmol) are dissolved in absolute ethanol (10.0 mL), concentrated hydrochloric acid (2-3 drops) is added as a catalyst, and the mixture is heated and refluxed for 4-5.5 hours. Light yellow solid is separated out in the reaction process, and the thin layer chromatography shows that the reaction is complete and is filtered. The solubility of the product is poor, and the target product with higher purity can be obtained by washing with ethyl acetate.
TABLE 2 reaction time, yield and physical Properties for the preparation of target Compounds 1a-1aa
Figure BDA0003509581640000072
Figure BDA0003509581640000081
Characterization of target Compounds
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N-phenylsemicarbazone (1 a). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.15 (s, 1H, ph-NH-pyrimidine ring), 9.83 (s, 1H, =N-NH-CO), 9.24 (s, 1H, CO-NH-Ph "), 8.61 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.87 (d, J=5.1 Hz,1H, pyrimidine ring CH 5 ),7.78–7.67(m,2H,Ph’H),7.59(dd,J＝10.4,4.2Hz,1H,Ph’H),7.54(d,J＝7.9Hz,2H,Ph”H 2,6 ),7.44(t,J＝7.0Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 ),7.31(t,J＝7.9Hz,2H,Ph”H 3,5 ),7.03(t,J＝7.4Hz,1H,Ph”H 4 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.74,159.48,159.04,152.02,145.26,143.96,139.15,133.15,132.95,131.81,131.57,130.27,129.20,128.45,123.35,120.10,119.99,118.29,109.27,102.48ppm；MS(ESI+)490(M+Na) + ；HPLC:t R ＝17.60min,98.5％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-methylphenyl) semicarbazone (1 b). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.15 (s, 1H, ph-NH-pyrimidine ring), 9.78 (s, 1H, =N-NH-CO), 9.16 (s, 1H, CO-NH-Ph "), 8.61 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.88 (d, J=5.1 Hz,1H, pyrimidine ring CH 5 ),7.74–7.68(m,2H,Ph’H),7.59(t,J＝7.3Hz,1H,Ph’H),7.43(t,J＝9.0Hz,5H,PhH 3,5 +Ph’H+Ph”H 2,6 ),7.37(d,J＝8.8Hz,2H,PhH 2,6 ),7.11(d,J＝8.3Hz,2H,Ph”H 3,5 ),1.98(s,3H,CH 3 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.76,159.47,159.03,152.06,145.27,143.57,136.55,133.15,132.95,132.29,131.79,131.59,130.27,129.59,128.44,120.22,120.00,118.29,109.27,102.46,20.88ppm；MS(ESI+)504(M+Na) + ；HRMS(ESI+):m/z＝482.1495,484.1470[M+H] + ；calcd.482.1496,484.1467for C 26 H 21 ClN 7 O+H；HPLC:t R ＝18.60min,99.2％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-cyanophenyl) semicarbazone (1 c). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.17 (s, 1H, ph-NH-pyrimidine ring), 10.08 (s, 1H, =N-NH-CO), 9.67 (s, 1H, CO-NH-Ph "), 8.63 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.86 (d, j=4.6 hz,1h, pyrimidine ring CH 5 ),7.74–7.68(m,6H,Ph’H,Ph”H 2,3,5,6 ),7.60(t,J＝6.7Hz,1H,Ph’H),7.43(t,J＝7.0Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.55,159.48,159.18,151.85,145.21,145.05,143.72,133.68,132.96,131.90,131.55,131.47,130.28,128.44,119.98,119.73,119.65,118.30,109.38,104.83,102.53ppm。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-nitrophenyl) semicarbazone (1 d) 1H NMR (400 MHz, DMSO-d) 6 ) Delta 10.23 (s, 1H, ph-NH-pyrimidine ring), 10.20 (s, 1H, =N-NH-CO), 10.13 (s, 1H, CO-NH-Ph "), 8.63 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ),8.21(d,J＝9.1Hz,2H,Ph”H 2,6 ) 7.82 (d, j=8.9hz, 3H, pyrimidine ring CH5, ph "H 3,5 ),7.71(q,J＝7.9Hz,2H,Ph’H),7.59(t,J＝7.3Hz,1H,Ph’H),7.44(t,J＝8.6Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.64,159.38,159.06,151.80,145.93,145.32,145.16,142.19,133.04,132.96,131.92,131.54,131.40,130.29,128.45,125.45,119.97,119.06,118.34,109.36,102.57,56.49,19.03ppm；HRMS(ESI+):m/z＝513.1191,515.1165[M+H] + ；calcd.513.1190,515.1161for C 25 H 18 ClN 8 O 3 +H。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-methoxyphenyl) semicarbazone (1 e). 1 H NMR(400MHz,DMSO-d 6 )δ:10.14(s,1H,Ph-NH-pyrimidine ring), 9.78 (s, 1H, =n-NH-CO), 9.12 (s, 1H, CO-NH-Ph "), 8.60 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, J=5.0 Hz,1H, pyrimidine ring CH 5 ),7.77–7.64(m,2H,Ph’H),7.59(t,J＝7.4Hz,1H,Ph’H),7.43(dt,J＝7.6,5.1Hz,5H,PhH 3,5 +Ph’H+Ph”H 2,6 ),7.37(d,J＝8.9Hz,2H,PhH 2,6 ),6.89(d,J＝8.9Hz,2H,Ph”H 3,5 ),3.35(s,3H,CH 3 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.77,159.47,158.98,155.71,152.32,145.28,143.56,133.17,132.95,132.03,131.76,131.61,130.26,128.42,122.23,120.01,118.29,114.34,109.29,102.45,55.66ppm；MS(ESI+)520.5(M+Na) + ；HPLC:t R ＝17.35min,99.1％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-ethoxyphenyl) semicarbazone (1 f). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.14 (s, 1H, ph-NH-pyrimidine ring), 9.76 (s, 1H, =N-NH-CO), 9.11 (s, 1H, CO-NH-Ph "), 8.60 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, J=5.1 Hz,1H, pyrimidine ring CH 5 ),7.77–7.65(m,2H,Ph’H),7.59(t,J＝7.3Hz,1H,Ph’H),7.43(t,J＝9.8Hz,5H,PhH 3,5 ,Ph’H+Ph”H 2,6 ),7.37(d,J＝8.8Hz,2H,PhH 2,6 ),6.88(d,J＝8.9Hz,2H,Ph”H 3,5 ),3.98(q,J＝7.0Hz,2H,CH 2 ),1.30(t,J＝7.0Hz,3H,CH 3 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.78,159.47,158.98,154.98,152.31,145.28,143.53,133.17,132.95,131.93,131.76,131.61,130.26,128.48,122.23,120.00,118.28,114.87,109.29,102.45,63.58,15.19ppm；MS(ESI+)534.5(M+Na) + ；HRMS(ESI+)m/z:＝512.1602,514.1572[M+H] + ；calcd.512.1602,514.1572for C 27 H 23 ClN 7 O 2 +H；HPLC:t R ＝18.15min,98.5％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-hydroxyphenyl) semicarbazone (1 g). 1 H NMR(400MHz,DMSO-d 6 ) Delta: 10.13 (s, 1H, ph-NH-pyrimidine ring), 9.69 (s, 1H, =n-NH-CO), 9.19 (s, 1H, oh), 9.02 (s, 1H, CO-NH-Ph "), 8.60 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, J=5.2 Hz,1H, pyrimidine ring CH 5 ),7.76–7.64(m,2H,Ph’H),7.58(t,J＝7.3Hz,1H,Ph’H),7.42(dd,J＝12.1,8.3Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 ),7.28(d,J＝8.8Hz,2H,Ph”H 2,6 ),6.72(d,J＝8.8Hz,2H,Ph”H 3,5 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.81,159.46,158.95,153.89,152.36,145.28,143.33,133.17,132.95,131.61,131.58,130.41,130.25,128.48,122.65,120.00,118.28,115.58,109.27,102.43,56.51,19.03ppm；MS(ESI+)506(M+Na) + ；HRMS(ESI+)m/z:＝484.1283,486.1256[M+H] + ；calcd.484.1289,486.1259for C 25 H 19 ClN 7 O 2 +H；HPLC:t R ＝15.25min,98.1％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-fluorophenyl) semicarbazone (1 h). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.15 (s, 1H, ph-NH-pyrimidine ring), 9.87 (s, 1H, =N-NH-CO), 9.29 (s, 1H, CO-NH-Ph "), 8.62 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, J=4.8 Hz,1H, pyrimidine ring CH 5 ),7.71(dt,J＝15.3,7.8Hz,2H,Ph’H),7.57(dt,J＝8.9,6.3Hz,3H,Ph’H+Ph”H 2,6 ),7.43(t,J＝8.3Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.6Hz,2H,PhH 2,6 ),7.15(d,J＝8.8Hz,2H,Ph”H 3,5 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.70,159.47,159.04,157.30,145.26,143.99,135.45,133.15,132.95,131.79,131.58,130.26,128.41,122.27,120.00,118.29,115.82,115.60 109.33,102.47ppm；MS(ESI+)486.5(M+Na) + ；HPLC:t R ＝17.52min,98.5％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-chlorophenyl) semicarbazone (1 i). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.16 (s, 1H, ph-NH-pyrimidine ring), 9.93 (s, 1H, =n-NH-CO), 9.43 (s, 1H, CO-NH-Ph "), 8.62 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.88 (d, J=4.8 Hz,1H, pyrimidine ring CH 5 ),7.76-7.65(m,2H,Ph’H),7.58(d,J＝8.9Hz,3H,Ph’H+Ph”H 2,6 ),7.42(t,J＝9.1Hz,3H,PhH 3,5 +Ph’H),7.36(dd,J＝8.8,3.8Hz,4H,PhH 2,6 +Ph”H 3,5 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.69,159.42,159.03,152.09,145.22,144.29,138.19,133.11,132.95,131.52,130.26,129.05,128.43,126.96,121.63,119.99,118.31,109.33,102.50ppm；MS(ESI+)502.5(M+Na) + ；HRMS(ESI+)m/z:＝502.0957,504.0933[M+H] + ；calcd.502.0950,504.0920for C 25 H 18 Cl 2 N 7 O 2 +H；HPLC:t R ＝14.84min,98.2％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (4-bromophenyl) semicarbazone (1 j). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.16 (s, 1H, ph-NH-pyrimidine ring), 9.93 (s, 1H, =n-NH-CO), 9.36 (s, 1H, CO-NH-Ph "), 8.62 (d, j=5.1 hz,1H, pyrimidine ring CH) 6 ) 7.89 (d, J=4.2 Hz,1H, pyrimidine ring CH 5 ),7.71(q,J＝7.8Hz,2H,Ph’H),7.63-7.52(m,3H,Ph’H+Ph”H 2,6 ),7.49(d,J＝8.6Hz,2H,Ph”H 3,5 ),7.43(t,J＝7.4Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.5Hz,2H,PhH 2,6 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.65,159.48,159.09,152.06,145.25,144.30,138.62,133.13,132.95,131.95,131.83,131.55,130.26,128.43,122.04,119.99,118.29,114.95,109.35,102.49ppm；MS(ESI+)568.5(M+Na) + ；HRMS(ESI+)m/z:＝546.0439,548.0419[M+H]+；calcd.546.0445,548.0424for C 25 H 18 BrClN 7 O 2 +H；HPLC:t R ＝19.58min,99.2％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (3-methylphenyl) semicarbazone (1 k). Yield:48.2%; light yellow solid; mp 258.4-259.1 ℃; 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.16 (s, 1H, ph-NH-pyrimidine ring), 9.83 (s, 1H, =n-NH-CO), 9.21 (s, 1H, CO-NH-Ph "), 8.61 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.86 (d, J=5.0 Hz,1H, pyrimidine ring CH 5 ),7.77–7.66(m,2H,Ph’H),7.59(t,J＝7.3Hz,1H,Ph’H),7.43(t,J＝7.9Hz,3H,PhH 3,5 +Ph’H),7.36(t,J＝11Hz,4H,PhH 2,6 +Ph”H 2,6 ),7.18(t,J＝7.7Hz,1H,Ph”H 5 ),6.85(d,J＝7.4Hz,1H,Ph”H 4 ),2.28(s,3H,CH 3 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.76,159.44,159.02,151.99,145.25,143.88,139.06,138.40,132.96,131.81,131.56,130.27,129.05,128.45,124.07,120.56,120.00,118.30,117.22,109.25,102.47,21.66ppm；MS(ESI+)582(M+Na) + ；HRMS(ESI+)m/z:＝482.1493,484.1474[M+H] + ；calcd.482.1496,484.1467for C 26 H 21 ClN 7 O+H；HPLC:t R ＝14.55min,99.1％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (3-nitrophenyl) semicarbazone (1 l). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.17 (s, 1H, ph-NH-pyrimidine ring), 10.13 (s, 1H, =N-NH-CO), 9.78 (s, 1H, CO-NH-Ph "), 8.62 (s, 1H, ph" H) 2 ) 8.65 (d, J=5.2 Hz,1H, pyrimidine ring CH 6 ),7.94(s,2H,Ph”H 4,6 ) 7.89 (d, J=9.8 Hz,1H, pyrimidine ring CH 5 ),7.73(d,J＝7.4Hz,2H,Ph’H),7.60(t,J＝8.3Hz,2H,Ph’H+Ph”H 5 ),7.44(t,J＝8.8Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.57,159.48,159.14,152.34,148.49,145.23,144.84,140.59,133.12,132.94,131.84,131.54,130.47,130.26,128.40,119.98,118.30,117.75,114.13,109.46,102.50ppm；MS(ESI+)513.5(M+H) + ；HRMS(ESI+)m/z:＝513.1191,515.1165[M+H]+；calcd.513.1191,515.1164for C 25 H 18 ClN 8 O 3 +H；HPLC:t R ＝14.05min,98.1％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (3-hydroxyphenyl) semicarbazone (1 m). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.19 (s, 1H, ph-NH-pyrimidine ring), 9.82 (s, 1H, =N-NH-CO), 9.24 (s, 1H, CO-NH-Ph "), 8.60 (d, J=5.3 Hz,1H, pyrimidine ring CH) 6 ) 7.81 (d, J=5.1 Hz,1H, pyrimidine ring CH 5 ),7.77–7.63(m,2H,Ph’H),7.59(t,J＝7.3Hz,1H,Ph’H),7.43(t,J＝8.8Hz,3H,PhH 3,5 +Ph’H),7.37(t,J＝8.8Hz,2H,PhH 2,6 ),7.15-6.99(m,2H,Ph”H 2,5 ),6.89(d,J＝8.3Hz,1H,Ph”H 2 ),6.44(d,J＝9.5Hz,1H,Ph”H 4 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.92,159.29,158.81,158.19,151.73,145.18,143.82,140.22,133.11,132.96,131.83,131.60,131.50,130.28,129.85,128.47,119.98,118.35,110.43,106.87,102.55,56.50,19.03ppm。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (3-methoxyphenyl) semicarbazone (1N). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.19 (s, 1H, ph-NH-pyrimidine ring), 9.86 (s, 1H, =N-NH-CO), 9.36 (s, 1H, CO-NH-Ph "), 8.61 (d, J=5.3 Hz,1H, pyrimidine ring CH) 6 ) 7.82 (d, j=5.0 hz,1h, pyrimidine ring CH 5 ),7.78–7.65(m,2H,Ph’H),7.59(t,J＝7.4Hz,1H,Ph’H),7.43(t,J＝9.0Hz,3H,PhH 3,5 +Ph’H),7.38(t,J＝9.5Hz,2H,PhH 2,6 ),7.20(dd,J＝11.2,4.9Hz,2H,Ph”H 2,5 ),7.07(d,J＝8.0Hz,1H,Ph”H 6 ),6.61(d,J＝8.2Hz,1H,Ph”H 4 ),2.49(s,3H,CH 3 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.87,160.07,159.31,158.86,151.86,145.18,144.02,133.10,132.96,131.84,131.58,131.49,130.28,129.99,128.47,119.97,118.36,112.13,109.21,108.67,105.68,102.56,55.47ppm。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (3-fluorophenyl) semicarbazone (1 o). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.16 (s, 1H, ph-NH-pyrimidine ring), 9.95 (s, 1H, =n-NH-CO), 9.42 (s, 1H, CO-NH-Ph "), 8.63 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.88 (d, J=4.9 Hz,1H, pyrimidine ring CH 5 ),7.72(dt,J＝15.4,8.1Hz,2H,Ph’H),7.62-7.51(m,2H,Ph’H+Ph”H 2 ),7.47-7.40(m,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 ),7.34-7.28(m,2H,Ph”H 4,6 ),6.85(t,J＝7.7Hz,1H,Ph”H 5 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:163.19,162.62,161.52,159.48,159.12,152.01,145.24,144.48,141.10,140.98,133.12,132.96,131.85,131.53,130.71,130.27,128.44,119.99,118.29,115.71,109.34,106.55,102.49ppm；MS(ESI+)508(M+H) + ；HPLC:t R ＝16.05min,98.2％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (3-chlorophenyl) semicarbazone (1 p). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.16 (s, 1H, ph-NH-pyrimidine ring), 9.97 (s, 1H, =n-NH-CO), 9.40 (s, 1H, CO-NH-Ph "), 8.63 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.90 (d, j=5.0 hz,1h, pyrimidine ring CH 5 ),7.77–7.68(m,3H,Ph’H+Ph”H 2 ),7.70-7.41(m,4H,PhH 3,5 +Ph’H+Ph”H 2 ),7.59(t,J＝7.3Hz,1H,Ph’H),7.38-7.31(m,3H,PhH 2,6 +Ph”H 5,6 ),7.09(d,J＝7.8Hz,1H,Ph”H 4 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.61,159.48,159.12,152.10,145.24,144.48,140.74,133.53,133.13,132.96,131.84,131.55,130.81,130.26,128.42,122.99,119.99,119.50,118.52,118.29,102.49ppm；MS(ESI+)524.5(M+Na) + ；HRMS(ESI+):m/z:＝502.0939,504.0914[M+H]+；calcd.502.0950,504.0920for C 25 H 18 Cl 2 N 7 O 2 +H；HPLC:t R ＝19.20min,97.9％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (3-bromophenyl) semicarbazone (1 q). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.15 (s, 1H, ph-NH-pyrimidine ring), 9.98 (s, 1H, =N-NH-CO), 9.41 (s, 1H, CO-NH-Ph "), 8.63 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, J=5.0 Hz,2H, pyrimidine ring CH 5 +Ph”H 2 ),7.78–7.66(m,2H,Ph’H),7.59(t,J＝7.3Hz,1H,Ph’H),7.50(d,J＝8.0Hz,1H,Ph”H 6 ),7.43(t,J＝4.6Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.7Hz,2H,PhH 2,6 ),7.27(t,J＝8.0Hz,1H,Ph”H 5 ),7.21(d,J＝8.0Hz,1H,Ph”H 4 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.61,159.47,159.11,152.10,145.24,144.48,140.48,133.13,132.95,131.83,131.54,131.11,130.26,128.42,125.89,122.34,122.01,119.99,118.90,118.30,109.39,102.50ppm；MS(ESI+)568.5(M+Na) + ；HRMS(ESI+)m/z:＝546.0434,548.0412[M+H]+；calcd.546.0445,548.0424for C 25 H 18 BrClN 7 O 2 +H；HPLC:t R ＝16.18min,98.3％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (2-methylphenyl) amino-acetal (1 r). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.17 (s, 1H, ph-NH-pyrimidine ring), 9.85 (s, 1H, =N-NH-CO), 9.27 (s, 1H, CO-NH-Ph "), 8.61 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.85 (d, j=5.1 hz,1h, pyrimidine ring CH 5 ),7.77–7.66(m,2H,Ph’H),7.59(t,J＝7.2Hz,1H,Ph’H),7.43(t,J＝9.5Hz,3H,PhH 3,5 +Ph’H),7.38-7.33(m,4H,PhH 2,6 +Ph”H 5,6 ),7.18(t,J＝7.7Hz,1H,Ph”H 4 ),6.84(d,J＝7.4Hz,1H,Ph”H 3 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.85,159.49,159.35,158.89,151.98,145.21,143.84,139.09,138.38,133.13,132.95,131.81,131.59,131.54,130.27,129.04,128.45,124.04,120.53,119.98,118.34,117.18,109.24,102.52,21.66ppm；MS(ESI+)m/z 482(M+H) + ；HPLC:t R ＝14.56min,98.6％。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-one-N- (2-hydroxyphenyl) semicarbazone (1 s). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.35 (s, 1H, ph-NH-pyrimidine ring), 10.21 (s, 1H, =N-NH-CO), 8.94 (s, 1H, CO-NH-Ph "), 8.60 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 8.00 (d, J=7.9 Hz,1H, pyrimidine ring CH 5 ),7.74–7.67(m,2H,Ph’H),7.59(dd,J＝13.7,6.3Hz,2H,Ph’H+Ph”H 6 ),7.43(t,J＝7.6Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 ),6.90(d,J＝7.8Hz,1H,Ph”H 3 ),6.84(t,J＝7.5Hz,1H,Ph”H 4 ),6.76(t,J＝7.6Hz,1H,Ph”H 5 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:163.10,159.34,158.80,151.87,146.97,145.15,143.52,133.14,132.97,131.78,131.72,130.18,128.36,127.16,123.32,119.98,119.87,119.58,118.38,115.16,108.57,102.59ppm。
2-chlorophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (2-methoxyphenyl) semicarbazone (1 t). Delta. 10.35 (s, 1H, ph-NH-pyrimidine ring), 10.21 (s, 1H, =N-NH-CO), 8.94 (s, 1H, CO-NH-Ph "), 8.61 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.83 (d, j=4.9hz, 1h, pyrimidine ring CH) 5 ),7.75–7.68(m,2H,Ph’H),7.59(t,J＝7.2Hz,1H,Ph’H),7.43(t,J＝9.2Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.6Hz,2H,PhH 2,6 ),7.21-7.18(m,2H,Ph”H 4,5 ),7.07(d,J＝8.0Hz,1H,Ph”H 6 ),6.60(d,J＝8.0Hz,1H,Ph”H 3 ),2.49(s,3H,CH 3 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.88,160.06,159.30,158.83,151.85,145.17,144.00,140.42,133.11,132.96,131.84,131.59,131.50,130.28,129.99,128.47,119.98,118.36,112.13,109.20,108.66,105.67,102.56,55.47ppm。
2-bromophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (4-bromophenyl) semicarbazone (1 u). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.17 (s, 1H, ph-NH-pyrimidine ring), 9.96 (s, 1H, =N-NH-CO), 9.51 (s, 1H, CO-NH-Ph "), 8.64 (d, J=4.8 Hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, j=4.2 hz,3h, pyrimidine ring CH 5 +Ph”H 2,6 ),7.63(s,2H,Ph’H),7.51(d,J＝7.3Hz,1H,Ph’H),7.45(d,J＝8.0Hz,3H,PhH 3,5 +Ph’H),7.38(d,J＝7.9Hz,2H,PhH 2,6 ),7.28(d,J＝7.8Hz,2H,Ph”H 3,5 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.53,159.41,159.06,145.67,145.23,145.23,140.89,133.68,133.46,133.26,132.92,131.92,131.62,131.10,128.84,123.08,122.00,119.97,119.05,118.30,109.42,102.46ppm。
2-bromophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (4-chlorophenyl) semicarbazone (1 v). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.18 (s, 1H, ph-NH-pyrimidine ring), 9.93 (s, 1H, =n-NH-CO), 9.52 (s, 1H, CO-NH-Ph "), 8.62 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.88 (d, J=4.8 Hz,1H, pyrimidine ring CH 5 ),7.76-7.65(m,2H,Ph’H),7.58(d,J＝8.9Hz,3H,Ph’H+Ph”H 2,6 ),7.42(t,J＝9.1Hz,3H,PhH 3,5 +Ph’H),7.36(dd,J＝8.8,3.8Hz,4H,PhH 2,6 +Ph”H 3,5 )ppm； 13 C NMR(100MHz,DMSO-d 6 )δ:162.59,159.32,158.95,152.02,145.41,145.21,138.21,133.68,133.32,132.92,131.90,131.63,129.03,128.91,126.85,123.02,121.45,119.85,118.32,102.49ppm。
2-bromophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (4-fluorophenyl) semicarbazone (1 w). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.16 (s, 1H, ph-NH-pyrimidine ring), 9.85 (s, 1H, =n-NH-CO), 9.36 (s, 1H, CO-NH-Ph "), 8.63 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, j=3.5 hz,2h, pyrimidine ring CH 5 +Ph’H),7.65–7.62(m,2H,Ph’H),7.57(dd,J＝8.8,4.9Hz,2H,Ph’H),7.45(d,J＝8.8Hz,2H,PhH 3,5 ),7.38(d,J＝8.8Hz,3H,PhH 2,6 +Ph’H),7.19-7.10(m,2H,Ph” 3,5 )ppm。
2-bromophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (3-nitrophenyl) semicarbazone (1X). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.19 (s, 1H, ph-NH-pyrimidine ring), 10.11 (s, 1H, =N-NH-CO), 9.90 (s, 1H, CO-NH-Ph "), 8.66 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ),8.63(s,1H,Ph”H 2 ),7.95(d,J＝6.6Hz,2H,Ph”H 4,6 ) 7.92-7.87 (m, 2H, pyrimidine ring CH) 5 +Ph’H),7.64-7.59(m,3H,Ph’H),7.45(t,J＝8.8Hz,2H,PhH 3,5 ),7.40(dd,J＝13.7,5.8Hz,3H,PhH 2,6 +Ph’H)ppm。
2-bromophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (4-cyanophenyl) semicarbazone (1 y). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.16 (s, 1H, ph-NH-pyrimidine ring), 9.85 (s, 1H, =n-NH-CO), 9.36 (s, 1H, CO-NH-Ph "), 8.63 (d, j=5.2 hz,1H, pyrimidine ring CH) 6 ) 7.86 (d, j=4.6 hz,1h, pyrimidine ring CH 5 ),7.74–7.68(m,6H,Ph’H,Ph”H 2,3,5,6 ),7.60(t,J＝6.7Hz,1H,Ph’H),7.43(t,J＝7.0Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.8Hz,2H,PhH 2,6 )ppm。
2-bromophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (3-bromophenyl) semicarbazone (1 z). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.17 (s, 1H, ph-NH-pyrimidine ring), 9.95 (s, 1H, =N-NH-CO), 9.46 (s, 1H, CO-NH-Ph "), 8.64 (d, J=5.2 Hz,1H, pyrimidine ring CH) 6 ) 7.91 (d, J=5.0 Hz,2H, pyrimidine ring CH 5 +Ph”H 2 ),7.78–7.66(m,2H,Ph’H),7.59(t,J＝7.3Hz,1H,Ph’H),7.50(d,J＝8.0Hz,1H,Ph”H 6 ),7.43(t,J＝4.6Hz,3H,PhH 3,5 +Ph’H),7.37(d,J＝8.7Hz,2H,PhH 2,6 ),7.27(t,J＝8.0Hz,1H,Ph”H 5 ),7.21(d,J＝8.0Hz,1H,Ph”H 4 )ppm.
2-bromophenyl-2- (4-cyanophenylamino) -pyrimidin-4-yl-ketone-N- (3-methoxyphenyl) semicarbazone (1 aa). 1 H NMR(400MHz,DMSO-d 6 ) Delta 10.17 (s, 1H, ph-NH-pyrimidine ring), 9.95 (s, 1H, =N-NH-CO), 9.46 (s, 1H, CO-NH-Ph "), 8.64 (d, J=5.3 Hz,1H, pyrimidine ring CH) 6 ) 7.90 (d, j=5.0 hz,1h, pyrimidine ring CH 5 ),7.78–7.65(m,2H,Ph’H),7.59(t,J＝7.4Hz,1H,Ph’H),7.43(t,J＝9.0Hz,3H,PhH 3,5 +Ph’H),7.38(t,J＝9.5Hz,2H,PhH 2,6 ),7.20(dd,J＝11.2,4.9Hz,2H,Ph”H 2,5 ),7.07(d,J＝8.0Hz,1H,Ph”H 6 ),6.61(d,J＝8.2Hz,1H,Ph”H 4 ),2.49(s,3H,OCH 3 )ppm。
Example 3: anti-HIV biological Activity test
The in vitro cell level anti-HIV virus activity test mainly comprises: inhibitory Activity and cytotoxicity against HIV-infected MT-4 cells. The method comprises the following steps: the protection of HIV-mutagenized cytopathy by the drug was assayed by MTT method in HIV-infected MT-4 cells at various times during HIV infection, and the half-effective concentration EC was calculated at the concentration required to protect 50% of the cells from HIV-induced cytopathy 50 Toxicity assays were performed in parallel with anti-HIV activity assays, also in MT-4 cell culture, using MTT to determine the concentration of cytopathic 50% of uninfected cells (CC 50 ) And calculates a selectivity index si=cc 50 /EC 50 。
Materials and methods:
the anti-HIV activity of each compound is monitored by the efficiency of the drug's inhibition of HIV-induced cytopathic effects in the cells. Cell culture was performed using MT-4 cells. The virus strains used were: HIV-1 strain IIIB and HIV-2 strain ROD.
The specific operation is as follows: dissolving the compound in DMSO or water, diluting with phosphate buffer saline solution, and concentrating 3×10 5 MT-4 cells were pre-incubated with 100. Mu.L of each compound at various concentrations for 1h at 37℃and then 100. Mu.L of the appropriate viral dilutions were added to the compounds and the cells were incubated for 1h at 37 ℃. After three washes, the cells were resuspended in culture medium with or without compound, respectively. The cells were then exposed to 5% CO 2 The culture was continued for another 7 days at 37℃in the atmosphere, and the supplementary medium was replaced with medium with or without compound on the third day after infection. The procedure was repeated twice for each broth condition. Cytopathic effects on viruses were monitored daily with a reverse optical microscope. Typically, the viral dilutions used in this experiment often lead to cytopathic effects the fifth day after viral infection. Drug inhibition concentration is used for inhibiting virus cells by drugThe concentration at which the pathological effect produced 50% inhibition without direct toxicity to the cells (CC 50 ) And (3) representing. It is emphasized that when compounds are poorly water soluble and DMSO is required to be dissolved, the specific DMSO concentration is typically less than 10% relative to water (DMSO final concentration in MT-4 cell culture medium is less than 2%). Because DMSO can affect the antiviral activity of the test compounds, antiviral activity in solutions containing the same concentration of DMSO should also be run in parallel versus blank experiments. In addition, the final DMSO concentration (1/1000) was far lower than that required for HIV-1 replication in T cells.
The results of the inhibitory activity of some target compounds against HIV using the marketed drugs Nevirapine (NVP), efavirenz (EFV) and itravirenz (ETV) as controls are summarized in tables 3-4. Experimental results show that the compounds contained in the chemical general formula generally have stronger anti-HIV-1 virus activity, smaller cytotoxicity and higher selectivity index. The active cytotoxicity results of DAPY-SC compounds against HIV-1IIIB, mutant strain RES056 and HIV-2 strain ROD are shown in Table 3.
TABLE 3 anti-HIV Activity and cytotoxicity of target Compounds
Figure BDA0003509581640000161
Figure BDA0003509581640000171
As shown in Table 3, EC of 20 target compounds against wild-type HIV-1 50 The value is in the range of 0.0329-1.1538 mu M, has low micromolar inhibition activity on HIV-1 wild-type strains, and the SI value is in the range of 22-3712, so that the inhibition activity is reduced compared with a lead compound. Wherein the activity of the target compound 1h is optimal (r=4-F, EC 50 Has a value of 0.0329 mu M and an SI value of 3712), which is stronger than the NEV (EC) 50 A value of 0.114. Mu.M, SI value of 132), DEV (EC 50 A value of 0.0366. Mu.M, an SI value of 1200) and ddI (EC 50 The value was 19.96. Mu.M and the SI value was 11). In addition, 1a (H,EC 50 a value of 0.0449. Mu.M, SI value of 1575), 1c (4-CN, EC 50 0.0650. Mu.M and 519 SI), 1n (3-OCH) 3 ,EC 50 A value of 0.0402. Mu.M, an SI value of 213), 1p (3-Cl, EC 50 Values of 0.0737. Mu.M, SI value 1729) and 1s (2-OH, EC 50 0.0372. Mu.M and 135 SI) are also more potent than the reference drugs NEV and ddI. While EC of intermediate 13a 50 The value was 0.0111. Mu.M, SI was 5548, which is stronger than the reference drugs NEV, DEV and ddI, and the activity was stronger than 20 target compounds.
Part of the compounds showed weak inhibitory activity against double mutant strain RES056, including 1f (4-OC 2 H 5 ,EC 50 A value of 5.12. Mu.M), 1g (4-OH, EC 50 Value 4.92. Mu.M), 1M (3-OH, EC 50 A value of 5.21. Mu.M), 1n (3-OCH) 3 ,EC 50 Value 8.01. Mu.M), 1s (2-OH, EC 50 Values of 5.31. Mu.M) and 1t (2-OCH 3 ,EC 50 The value was 8.49. Mu.M), which is superior to the reference drug NEV (EC) 50 Values of 15.04. Mu.M) and DEV (EC 50 Values 43.86. Mu.M).
In addition, some compounds exhibit some activity against HIV-2ROD, including 1f (4-OC 2 H 5 ,EC 50 Value 6.84. Mu.M), 1g (4-OH, EC 50 Value 5.14. Mu.M), 1M (3-OH, EC 50 A value of 5.97. Mu.M), 1n (3-OCH) 3 ,EC 50 Value 8.99. Mu.M), 1s (2-OH, EC 50 Values of 4.67. Mu.M) and 1t (2-OCH 3 ,EC 50 The value was 9.94. Mu.M). Notably, this is consistent with the inhibitory activity on double mutant strain RES 056.
Example 4 test of target Compounds against HIV-1RT Activity at enzyme level
To confirm that the target of the target compound is HIV-1RT, the present patent conducted enzyme level inhibition activity tests on the target compound. Reverse transcriptase activity assay kit (EnzCheck Reverse Transcriptase Assay kit) for experimental materials was purchased from Invitrogen corporation. Positive controls Nevirapine (NEV) and Efavirenz (EFV).
(1) Test principle and method
Recombinant wild-type reverse transcriptase P66/P51 HIV-1RT was expressed and purified according to the methods reported in the literature and reverse transcriptase activity assay was performed using a commercial reverse transcriptase activity assay kit (EnzCheck Reverse Transcriptase Assay kit), all steps being performed strictly according to the instructions of the test kit. The function principle of the detection box is that the fluorescence signal of the dye PicoGreen is obviously enhanced when the dye PicoGreen is combined with double-stranded DNA or RNA/DNA hybrid double-stranded, so that the quantitative effect on double-stranded nucleic acid is achieved. And even in the presence of very high dyes: the single stranded nucleic acids have only slight fluorescent signals in base pairing.
(2) Experimental procedure
Oligo (dT) with 350 bases Poly (rA) as template 16 As primers, annealing at room temperature for 60 minutes at a molar ratio of 1:1.2: mu.L of a polymerization buffer (containing 60mM Tri-HCl,60mM KCl,8mM MgCl) containing 52ng of the RNA/DNA mixture was added to each well in a 96-well plate 2 13mM DTT,100mM dTTP,pH 8.1); mu.L of RT enzyme solution was diluted to the appropriate concentration with enzyme dilution (containing 50mM Tri-HCl,20%glycerol,2mM DTT,pH 7.6); the reaction was incubated at 25℃for 40 minutes, quenched by the addition of 15mM EDTA, and then hybridized duplex was detected by the addition of PicoGreen dye. Absorbance values at an emission wavelength of 523nm at an excitation wavelength of 490nm were determined by means of an enzyme-labeled instrument. To test the anti-reverse transcriptase activity of the compounds, 1 μl of compound solution in DMSO was pre-added to each well prior to adding the reverse transcriptase solution. The samples to be tested were dissolved in DMSO to the appropriate concentration and then diluted 5-fold with DMSO, 8 dilutions each. While control wells were added 1 μl of DMSO without compound. The results are expressed as relative fluorescence values, i.e. fluorescence intensity of wells containing compound/fluorescence intensity of wells not containing compound.
(3) Results and discussion
The results of the inhibitory activity of the target compounds against wild-type HIV-RT are shown in Table 4.
TABLE 4 inhibitory Activity of target Compounds against HIV-1RT
Figure BDA0003509581640000181
Figure BDA0003509581640000191
As shown in Table 4, 20 target compounds and 13a each have strong inhibitory activity against HIV-1RT, IC of the target compound 50 The value is 0.438-10.14 mu M. Of these, 1h had the strongest inhibitory activity (IC 50 The value is 0.438 mu M), which is superior to the NEV (IC) reference medicine 50 The value was 0.971. Mu.M). In addition, 1a (IC 50 Value 0.522. Mu.M), 1l (IC 50 0.970. Mu.M), 1n (IC 50 A value of 0.833. Mu.M), 1q (IC) 50 Values of 0.955. Mu.M) and 1s (IC 50 A value of 0.594 μm) was also stronger than the reference drug NEV. The target compound is HIV-1RT.
In summary, the results of experiments on both cellular and enzymatic levels indicate that the compounds comprised in the chemical formula generally have a strong anti-HIV-1 virus activity, a small cytotoxicity and a high selectivity index.
Molecular Docking (dock) analysis between compounds and target RT
In order to obtain a potential active conformation of the molecule, a Surflex-Dock module in a Sybyl molecular simulation software package is used, TMC125/HIV-1 complex 3MEC is selected according to the research result of 2D-QSAR, and the enzyme protein is dehydrated and hydrogenated after ligand is extracted. The Gasteig-Huckel charge was loaded after conformational optimization using the steepest energy gradient descent method under the Tripos force field. The optimized molecules were mock-docked with the prepared receptor proteins using the extracted TMC125 conformation as a reference. Parameters of the docking results are in the appendix. Comparing with the Total Score (TS) value, 1ac is highest, and the TS values of 1q,1p,1j and 1k are also higher. The docking score value matches the activity prediction of the compound of interest using the QSAR model.
The interaction of the ligand with the reverse transcriptase was observed, taking as an example the binding pattern of 1q,1p,1k with a strong predicted activity and 1d and 3MEC with a poor predicted result, the binding pattern is shown by the analysis of fig. 2.
The conformation of 1q and 1p with stronger prediction activity in NNIBP of RT is similar, and the ligand is in U-shaped conformation, and enters into hydrophobic pocket formed by amino acids of Tyr181, tyr188, trp229 and the like to form hydrophobic interaction, and the left wing aromatic ring and benzene rings of Tyr181 and Tyr188 are in the same plane to form pi-pi stacking interaction. Meanwhile, the right wing aromatic ring linker NH forms hydrogen bonds with Lys 101. NH on 1q semicarbazone can also form hydrogen bonds with Ile180, binding to enzymes more tightly.
While the conformation of 1k and the poorly predicted 1d in the hydrophobic binding pocket of the enzyme is different from the classical U-shaped conformation. The position of the left wing aromatic ring of 1k is exchanged with 1q and 1p compared with the aromatic ring of semicarbazone, and the benzene ring on semicarbazone enters a hydrophobic pocket to form pi-pi stacking interaction. 1d is far away from hydrophobic pockets formed by amino acids like Tyr181, tyr188 and Trp229, and cannot form pi-pi stacking interactions and hydrophobic interactions with enzymes, while 1d can also form hydrogen bonds with Lys101, but its affinity to enzymes is greatly reduced by 1p,1q and 1k, and it is seen that interactions with hydrophobic pockets are important for ligand binding to HIV-1RT.
The invention is not limited to the examples described above.
Reference is made to:
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2.Meng,G.；Chen,F.-E.；De Clercq,E.；Balzarini,J.；Pannecouque,C.,Nonnucleoside HIV-1reverse transcriptase inhibitors:part I.Synthesis and structure-activity relationship of1-alkoxymethyl-5-alkyl-6-naphthylmethyl uracils as HEPT analogues.Chemical and Pharmaceutical Bulletin 2003,51(7),779-789.
3.Jin,K.；Sang,Y.；De Clercq,E.；Pannecouque,C.；Meng,G.,Design and synthesis of a novel series of non-nucleoside HIV-1inhibitors bearing pyrimidine and N-substituted aromatic piperazine.Bioorganic&Medicinal Chemistry Letters 2018,28,3491–3495.
4.Jin,K.；Sang,Y.；Han,S.；Clercq,E.D.；Pannecouque,C.；Meng,G.；Chen,F.,Synthesis and biological evaluation of dihydroquinazoline-2-amines as potentnon-nucleoside reverse transcriptase inhibitors of wild-type and mutant HIV-1strains.European Journal of Medicinal Chemistry 2019,176,11-20.
5.Jin,K.；Liu,M.；Zhuang,C.；De Clercq,E.；Pannecouque,C.；Meng,G.；Chen,F.,Improving the positional adaptability:structure-based design of biphenyl-substituted diaryltriazines as novel non-nucleoside HIV-1reverse transcriptase inhibitors.Acta Pharm.Sin.B 2020,10(2),344-357.
6.Xiao,T.；Tang,J.-F.；Meng,G.；Pannecouque,C.；Zhu,Y.-Y.；Liu,G.-Y.；Xu,Z.-Q.；Wu,F.-S.；Gu,S.-X.；Chen,F.-E.,Indazolyl-substituted piperidin-4-yl-aminopyrimidines as HIV-1NNRTIs:Design,synthesis and biological activities.European Journal of Medicinal Chemistry 2020,186(2020)111864(186),111864.
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Claims (7)

1. an HIV-1 reverse transcriptase inhibitor is characterized in that the inhibitor is a diaryl pyrimidine derivative containing substituted aryl urea imino, and has a structural formula shown in the following formula (I):
Figure FDA0004145653190000011
wherein R is independently selected from hydrogen, methyl, cyano, nitro, methoxy, ethoxy, hydroxy and halogen, and the substitution position can be ortho, para or meta; x is halogen.
2. The HIV-1 reverse transcriptase inhibitor of claim 1, wherein the number of diaryl pyrimidine derivatives containing a substituted aryl urea imino group is 27: 1a,1b, …,1z,1aa, which correspond to R, X as follows:
Figure FDA0004145653190000012
3. the HIV-1 reverse transcriptase inhibitor of claim 1, further comprising a pharmaceutically acceptable salt of a diaryl pyrimidine derivative containing a substituted aryl urea imino group, said pharmaceutically acceptable salt being a hydrochloride, hydrobromide, sulfate, phosphate, acetate, mesylate, p-toluenesulfonate, tartrate, citrate, fumarate or malate salt.
4. The method of synthesizing an HIV-1 reverse transcriptase inhibitor of claim 1, wherein the specific synthetic route is as follows:
Figure FDA0004145653190000021
the specific steps of the synthesis are as follows;
firstly, thiouracil (5) is used as a starting material, methyl iodide (6) is used as a methylation reagent, and under the action of sodium hydroxide, the reaction is carried out for 20 to 24 hours at room temperature, and S-alkylation reaction is carried out, so that a white solid with high purity, namely 2-methylthiopyrimidine-4-ketone (7), is obtained; wherein the amount of thiouracil (5) is 1.5-1.6 equiv, the amount of methyl iodide (6) is 1.0-1.1 equiv, the molar concentration of sodium hydroxide is 1.20-1.35M, and the amount is 1.0-1.1 equiv;
secondly, reacting 2-methylthiopyrimidine-4-ketone (7) with excessive 4-cyanoaniline (8) in a molten state at 180-185 ℃ for 10-18 h under the condition of no solvent, and performing aftertreatment after acetonitrile dissolution to obtain yellow solid 2- (4-cyanoanilino) pyrimidine-4-ketone (9); wherein the amount of 2-methylthiopyrimidine-4-ketone (7) is 1.0-1.1 equiv., and the amount of 4-cyanoaniline (8) is 2.5-3.0 equiv.;
dissolving and refluxing 2- (4-cyanoanilino) pyrimidine-4-ketone (9) in excessive phosphorus oxychloride, performing chlorination reaction on hydroxyl at C-4 position of pyrimidine heterocycle, performing aftertreatment, dissolving in a proper amount of cold water, neutralizing to be neutral by sodium hydroxide to obtain yellow precipitate, and filtering and drying to obtain yellow solid 2- (4-cyanoanilino) -4-chloro-pyrimidine (10); wherein the amount of 2- (4-cyanoanilino) pyrimidin-4-one (9) is 1.0 to 1.1equiv., and the amount of phosphorus oxychloride is 10.0 to 11.0equiv.;
nucleophilic substitution reaction of 2- (4-cyanoanilino) -4-chloro-pyrimidine (10) and 2-halogeno-benzyl cyanide 11a,11b in dried N, N-dimethylformamide under the action of 60% sodium hydride, and no water and oxygen condition to obtain unstable intermediate Cyan-CH 2 DAPYs, the intermediate being of formula 12a,12b; wherein the amount of 2- (4-cyanoanilino) -4-chloro-pyrimidine (10) is 1.0-1.1 equiv., the amount of 2-halogenated benzyl cyanide 11a,11b is 1.5-1.6 equiv., the amount of sodium hydride is 2.0-2.4 equiv., and the amount of N, N-dimethylformamide is 2.5 equiv-3.0 mL;
(V) due to the above intermediate Cyan-CH 2 The DAPYs is unstable, the intermediate structural formulas 12a and 12b are removed from nitrogen protection after reaction, and are placed in air to react for 48 to 72 hours at room temperature, and are oxidized slowly to obtain an intermediate Oxo-CH 2 DAPYs, post-treatment and column chromatography separation to obtain pure 2-halophenyl 2- (4-cyanophenylamino) -pyrimidinone 13a,13b;
step six, simultaneously carrying out the steps one to five, taking various substituted anilines as starting materials, and carrying out the preparation of various substituted semicarbazide 18a-18t through two steps of reactions in parallel; the specific method comprises dissolving various substituted anilines 14a-14t in tetrahydrofuran, and simultaneously dissolving NaHCO 3 Dissolving NaHCO in water 3 Mixing the aqueous solution with tetrahydrofuran solution of substituted aniline, placing the mixture in an ice bath, and adding phenyl chloroformate (15) after the temperature is stabilized at 0-5 ℃; the reaction speed is extremely high, and the reaction can be completed after the reactants are added; extracting with ethyl acetate, and rotary evaporating to obtain various stable substituted phenyl carbamate intermediates 16a-16t; dissolving the substituted phenyl carbamate intermediate 16a-16t in acetonitrile, adding 80% hydrazine hydrate (17), and reacting for 1-3 h at ultrasonic room temperature to obtain corresponding substituted semicarbazide 18a-18t; wherein, the various substituted anilines 14a-14t are 1.0 to 1.1equiv., naHCO 3 1.2 to 1.4equiv., and 1.2 to 1.4equiv. of phenyl chloroformate (15); the substituted phenyl carbamate intermediate 16a-16t is 1.0-1.2 equiv., and the hydrazine hydrate (17) is 2.5-3.0 equiv.;
(seventh) finally, the intermediate Oxo-CH 2 DAPYs and various substituted semicarbazide 18a-18t are heated, refluxed and dehydrated in ethanol for 4-5 hours under the condition of taking hydrochloric acid as a catalyst, so as to obtain corresponding target compounds 1a,1b, …,1z,1aa; wherein, the intermediate Oxo-CH 2 DAPYs is 1.0 to 1.1equiv, and the various substituted semicarbazides 18a-18t are 1.0 to 1.1 equiv;
in the sixth step, the substituted aniline 14a-14t, the substituted phenyl carbamate intermediate 16a-16t, the substituted semicarbazide 18a-18t, and the target objects 1a,1b, …,1z,1aa obtained in the seventh step, corresponding X, R are specifically listed as follows:
Figure FDA0004145653190000031
Figure FDA0004145653190000041
5. a pharmaceutical composition comprising an effective amount of any of the compounds of claim 1 and a pharmaceutically acceptable carrier.
6. The application of diaryl pyrimidine derivatives containing substituted aryl urea imino in preparation of HIV-1 reverse transcriptase inhibitors is provided, wherein the diaryl pyrimidine derivatives containing substituted aryl urea imino have a structural formula shown in the following formula (I):
Figure FDA0004145653190000042
wherein R is independently selected from hydrogen, methyl, cyano, nitro, methoxy, ethoxy, hydroxy, halogen, and the substitution position can be ortho, para or meta; x is halogen.
7. The use according to claim 6, for the preparation of a medicament for the prevention and treatment of aids.
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