4'-Fluoro-1,2,3-triazole riboside analogues, preparation methods thereof, and use thereof as anti-rabies virus inhibitors

Abstract

The present invention belongs to the field of medicinal chemistry and relates to a class of 4'-fluoro-1,2,3-triazole nucleoside analogs, their preparation methods, and their use as anti-rabies virus inhibitors. The structural formula of the triazole nucleoside analogs of the present invention is: #imgabs0#;R 1 、R 2 is benzyl, 4-methoxybenzyl, 4-bromobenzyl or 2-naphthylbenzyl; R 3 The present invention relates to a method for preparing a cyclopropyl group, a phenyl group, a 1-methyl-1-hydroxyethyl group, or a methoxycarbonyl group. Most reactions in the preparation method of the present invention are insensitive to air and water, and multi-step reactions can be continued without purification, greatly simplifying the operation, reducing the difficulty of synthesis, and being suitable for large-scale preparation. Such compounds have anti-rabies virus activity and can be used to prepare anti-rabies virus inhibitors.

Description

Technical Field

[0001] The present invention belongs to the field of medicinal chemistry and relates to a class of 4'-fluoro-1,2,3-triazole nucleoside analogs and a preparation method thereof, as well as the application of the compounds as anti-rabies virus inhibitors. Technical Background

[0002] Rabies is a neurological zoonotic disease caused by the rabies virus (RABV). Once clinical symptoms appear, it is almost 100% fatal. To date, there is no effective treatment to cure the disease after clinical symptoms appear. It is reported that approximately 60,000 people die from rabies each year worldwide.

[0003] Despite the availability of effective rabies prevention measures, such as human rabies vaccines and immunoglobulins, the disease remains a high risk in low-income areas, such as rural areas. To date, there are no effective drugs that can cure the disease after clinical symptoms appear. Therefore, the development of highly effective drugs specifically targeting rabies remains a hot topic in drug research and development.

[0004] Nucleoside drugs are an important class of antiviral drugs that play a key role in the treatment of infections caused by human immunodeficiency virus (HIV), hepatitis B virus (HBV), and hepatitis C virus (HCV). Their mechanism of action is to inhibit the activity of viral RNA polymerase, either by binding to the enzyme as a competitive inhibitor and inactivating it, or by incorporating into virally synthesized RNA sequences as a reaction substrate, preventing further elongation. This mechanism of action dictates that nucleoside drugs are more potent than other competitive inhibitors, directly preventing the reaction from proceeding rather than slowing it down. However, viral and cellular nucleic acid polymerases share certain similarities, which contributes to their cytotoxicity.

[0005] Ribovirin is a class of compounds in which the heterocyclic base in the nucleoside is replaced by a triazole heterocycle. The triazole heterocycle itself has been shown to have antiviral reverse transcriptase activity (J. Heterocycl. Chem. 1982, 19, 263). This class of compounds can be easily synthesized using a "click chemistry" method catalyzed by inexpensive and readily available copper salts (Bioorg. Med. Chem. 2008, 16, 8427). Ribovirin synthesized using this method has been shown to exhibit excellent activity against influenza A (H1N1) virus and HIV (Med. Chem. Res. 2014, 23, 1501). Summary of the Invention

[0006] Based on the above situation, the first object of the present invention is to provide a new class of 4'-fluoro-1,2,3-triazole riboside analogs with anti-rabies virus activity.

[0007] Another object of the present invention is to provide a large-scale preparation method of 4'-fluoro-1,2,3-triazole nucleoside analogs with readily available raw materials and low preparation difficulty.

[0008] The third object of the present invention is to provide the use of the above-mentioned 4'-fluoro-1,2,3-triazole riboside analogues for preparing anti-rabies virus inhibitors.

[0009] The molecular structure of the 4'-fluoro-1,2,3-triazole nucleoside analog of the present invention comprises the following features: (1) the 3' and 5' positions thereof are protected by alkoxy groups; and (2) the 4-position of the triazole is substituted by a functional group.

[0010] The general structural formula of the triazole riboside analogues of the present invention is shown below:

[0011]

[0012] R at the 3' or 5' position of ribavirin 1 or R 2 The substituent is a benzyl protecting group, which can be any one of benzyl, 4-methoxybenzyl, 4-bromobenzyl, and 2-naphthylbenzyl;

[0013] R at the 4-position of the triazole nucleoside base 3 The substituent is one of cyclopropyl, phenyl, 1-methyl-1-hydroxyethyl, and methoxycarbonyl.

[0014] The preparation method of the triazole riboside analog of the present invention comprises the following steps:

[0015] (1) Using the known compound 1 (Org. Chem. Front., 2022, 9, 2808–2814) as the starting material, the acetoxy group of 1 was activated under acidic conditions and glycosylated with trimethylsilyl azide to obtain the terminal azide compound 2;

[0016] (2) Removal of R from azide compound 2 under alkaline conditions 4 and acetoxy protecting group to obtain uronic acid 3;

[0017] (3) decarboxylating and fluorinating the carboxyl group of uronic acid 3 to obtain compound 4 or compound 5;

[0018] (4) The azide group of compound 4 or compound 5 undergoes a "click reaction" with an alkyne to obtain triazole nucleoside analogues 6 or 7.

[0019] The synthetic route of the present invention is as follows:

[0020]

[0021] Among them, R1 It is a benzyl protecting group, which can be any one of benzyl, 4-methoxybenzyl, 4-bromobenzyl, and 2-naphthylbenzyl, and is preferably benzyl.

[0022] R 2 It is a benzyl protecting group, which can be any one of benzyl, 4-methoxybenzyl, 4-bromobenzyl, and 2-naphthylbenzyl, and is preferably benzyl.

[0023] R 3 It can be an alkyl group, an aromatic group, an alkyl alcohol group, an alkoxycarbonyl group, an imine ester group, a cyano group or an amide group. 3 It is one of cyclopropyl, phenyl, 1-methyl-1-hydroxyethyl, and methoxycarbonyl.

[0024]

[0025] R 4 It is an alkyl protecting group, which can be any one of methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl and tert-butyl.

[0026] The specific preparation steps of the present invention are as follows:

[0027] Step 1: Preparation of compound 2:

[0028] In an organic solvent, compound 1 is reacted with trimethylsilyl azide at -78-100° C. under 1-100 atmospheres and for 30-3000 minutes under the catalysis of protonic acid or Lewis acid.

[0029] The organic solvent is a C1-C6 monohalogenated or polyhalogenated alkane or halogenated aromatic hydrocarbon solvent, preferably one or a mixture of dichloromethane, chloroform, 1,2-dichloroethane, chlorobenzene, trifluorotoluene.

[0030] The protonic acid is one or more of perchloric acid, tetrafluoroboric acid, tetrakis(pentafluorophenyl)boric acid, bis(trifluoromethanesulfonyl)imide, trifluoroacetic acid, formic acid, acetic acid, sulfuric acid-silica gel complex, perchloric acid-silica gel complex, trifluoromethanesulfonic acid-silica gel complex, trichloroacetic acid, trifluoromethanesulfonic acid, p-toluenesulfonic acid, and sulfuric acid.

[0031] The Lewis acid is one or a mixture of trimethylsilyl trifluoromethanesulfonate, triethylsilyl trifluoromethanesulfonate, tert-butyldimethyl trifluoromethanesulfonate, boron trifluoride ethyl etherate, zinc trifluoromethanesulfonate, scandium trifluoromethanesulfonate, lanthanum trifluoromethanesulfonate, ytterbium trifluoromethanesulfonate, indium trifluoromethanesulfonate, silver trifluoromethanesulfonate, silver tetrakis(pentafluorophenylborate), and tin tetrachloride.

[0032] The molar ratio of the compound 1, protonic acid or Lewis acid, and trimethylsilyl azide is 1:(0.1-100):(1-100); the preferred molar ratio is 1:0.4:2.

[0033] Step 2: Preparation of compound 3:

[0034] In a mixed solvent of water and an organic solvent, compound 2 is reacted at -78-100° C. for 30-3000 minutes under the action of a base.

[0035] The organic solvent is one or a mixture of methanol, ethanol, isopropanol and other C1-C6 alkyl alcohols, tetrahydrofuran, and dioxane; the preferred solvent is tetrahydrofuran.

[0036] The volume ratio of the mixed solvent is water: organic solvent is 1: (1-1000); the preferred volume ratio is 1:4.

[0037] The base is an inorganic base such as one or more of hydroxide, hydride, carbonate or ammonia; or an organic base such as one or more of potassium tert-butoxide, sodium tert-butoxide, lithium tert-butoxide, sodium methoxide, sodium ethoxide, methoxymagnesium, tetrabutylammonium fluoride.

[0038] The molar ratio of the compound 2 to the base is 1:(1-100); the preferred molar ratio is 1:4.

[0039] Step 3: Preparation of compound 4 or compound 5:

[0040] In a mixed solvent consisting of water and acetone, compound 3 reacts with a catalyst, an additive and a free radical acceptor at 1-100 atmospheric pressure and 0-100° C. for 10-3000 minutes.

[0041] The volume ratio of water to acetone in the mixed solvent is 1:(1-100), preferably 1:6.

[0042] The catalyst is a mixture of one or more of silver salt, iron salt, iridium salt, ruthenium salt, cerium salt and organic dye, and the base used is.

[0043] The additive is potassium fluoride dihydrate, potassium bromide, potassium iodide, etc., and the preferred additive is potassium fluoride dihydrate.

[0044] The free radical acceptor is 1-chloromethyl-4-fluoro-1,4-diazabicyclo[2,2,2]-octane bis(tetrafluoroborate) salt (Selectfluor).

[0045] The molar ratio of the compound 3, the catalyst, the additive and the free radical acceptor is 1:(0.01-100):(1-100):(1-100); the preferred molar ratio is 1:0.2:2:2.

[0046] Step 4: Preparation of compound 6 or compound 7:

[0047] In a mixed solvent consisting of an organic solvent and water, compound 4 or compound 5 reacts with alkyne, copper salt and sodium vitamin C respectively at 1-100 atmospheres and 0-100° C. for 30-3000 minutes.

[0048] The organic solvent is one or more of DMF, DMA, DMSO, methanol, ethanol, and tert-butanol, and the preferred solvent is DMF.

[0049] The alkyne is a terminal alkyne, preferably cyclopropylacetylene, phenylacetylene, 2-methyl-3-butyn-2-ol, or methyl propiolate.

[0050] The copper salt is a mixture of any one or more of copper sulfate, copper carbonate, and copper chloride, preferably copper sulfate.

[0051] The molar ratio of compound 4 or compound 5, alkyne, copper salt and sodium vitamin C is 1: (1-100): (0.1-100): (0.1-100); the preferred molar ratio is 1: 3.0: 0.4: 0.8.

[0052] The above 4'-fluoro-1,2,3-triazole riboside analogue is used for preparing anti-rabies virus inhibitors.

[0053] The advantages of the present invention are as follows: (1) the raw materials used can be obtained by simple conversion of commercially available diacetone glucose; (2) most of the reactions in this route are insensitive to oxygen and water, and multi-step reactions can be continued without purification, which reduces the difficulty of synthesis and is suitable for large-scale preparation; (3) the provided novel 4'-substituted 1,2,3-triazole nucleoside analogues have anti-rabies virus activity and can be used to treat rabies. In vivo pharmacokinetics show that the compounds can penetrate the blood-brain barrier and can be beneficial for solving diseases caused by rabies virus infection; (4) the activity studies of the provided 4'-substituted 1,2,3-triazole nucleoside analogues show that the IC 50 Below 10 μM and above 100 μM concentration, there is no significant inhibitory effect on cells, and the safety is relatively high; (5) The provided 4'-substituted 1,2,3-triazole nucleoside analogues effectively inhibit RABV infection by inhibiting viral polymerase to block viral replication, and are expected to become candidate drugs for anti-RABV. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] Figure 1 This is a graph showing the inhibition rate of different compounds on vesicular stomatitis virus at 50 μmol.

[0055] Figure 2 This is a graph showing that compound 7a inhibits RABV proliferation in vitro.

[0056] Figure A shows the cytotoxicity of 7a against BSR cells in the range of 0-120 μM, as assessed by the MTS assay. Figures B, C, D, and E used the following experimental methods: BSR cells were infected with RABV (MOI = 0.01) for 1 h at 37°C. The cell supernatant was then replaced with fresh culture medium containing 0 μM, 20 μM, 40 μM, 60 μM, or 80 μM, respectively. At 48 h, fixed cells were stained with FITC-RABV-P antibody, and nuclei were stained with DAPI (Figure B). Supernatants were collected for virus titration (Figure C). Figure D shows the RABV vRNA copy number assayed by RT-qPCR. Figure E shows the RABV-N protein assay by western blot, with β-actin as a control. Figure F shows the relative protein levels analyzed using Image-J. All results are the mean ± SD of three independent experiments. Significant differences were determined by one-way analysis of variance. Statistical differences are represented as follows: *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

[0057] Figure 3 The plasma drug concentration-time curves of mice after intravenous (3 mg / kg) and intramuscular (30 mg / kg) injection of compound 7a.

[0058] Figure 4 This is the tissue distribution diagram of mice after intramuscular injection of compound 7a (30 mg / kg). DETAILED DESCRIPTION

[0059] The present invention will be described in detail below with reference to the accompanying drawings and through specific synthesis examples.

[0060] Example 1:

[0061] Step 1: Preparation and data of compound 2:

[0062] To a 25 mL reaction flask equipped with a magnetic stirrer, compound 1 (250 mg, 0.53 mmol, 1.0 equiv) and trimethylsilyl azide (TMSN3, 141 μL, 1.06 mmol, 2.0 equiv) were added sequentially, and a dry dichloromethane solution (6 mL) was added for dissolution. Tin tetrachloride (25 μL, 0.212 mmol, 0.4 equiv) was slowly added under ice bath conditions. The reaction solution was stirred at room temperature for 1 h. After TLC, the reaction was complete and triethylamine was added to quench the reaction. The solvent was removed under reduced pressure and purified by column chromatography (petroleum ether:ethyl acetate = 7:1) to give a colorless syrup 2 (236 mg, 0.52 mmol, 98%).

[0063] The analysis data are as follows:

[0064] 1 H NMR(500MHz, CDCl3)δ:7.39–7.26(m,8H),7.25–7.20(m,2H),5.54(d,J=4.8Hz,1H),4.96(t,J=5.1Hz,1H),4.62(d,J=12.3Hz,1H),4.56( d,J=12.3Hz,1H),4.51(d,J=11.4Hz,1H),4.46(d,J=11.4Hz,1H),4.37(d,J=5.4Hz,1H),3.78(d,J=1.7Hz,2H),3.71(s,3H),2.04(s,3H);

[0065] 13 C NMR(125MHz, CDCl3)δ:170.1,169.3,137.6,137.2,128.6,128.5,128.1,1 27.9,127.9,127.9,93.2,90.1,79.1,75.4,74.8,74.0,71.9,52.6,20.7;

[0066] HRMS(ESI)m / z Calcd for C 23 H 29 O7N4[M+NH4] + 473.2031,found 473.2027.

[0067] Step 2: Preparation and data of compound 3:

[0068] To a 25 mL reaction flask equipped with a magnetic stirrer, compound 2 (69 mg, 0.152 mmol, 1.0 equiv) and lithium hydroxide monohydrate (25 mg, 0.606 mmol, 4.0 equiv) were added sequentially, and water / tetrahydrofuran (v:v = 1:4, 1.8 mL) was added for dissolution. The reaction was stirred at room temperature for 1 h, and then cationic resin (hydrogen form) was added to quench the reaction. The reaction was filtered, and the solvent was removed under reduced pressure. The mixture was separated and purified by column chromatography (petroleum ether: ethyl acetate = 3:1 → dichloromethane / ethyl acetate = 1.5:1) to give a colorless syrup 3 (51 mg, 0.13 mmol, 85%).

[0069] The analysis data are as follows:

[0070] 1 H NMR(500MHz, CDCl3)δ:7.4–7.24(m,10H),5.51(s,1H),4.90–4.53(m,5H),4.31(d,J=4.7Hz,1H),4.00(dd,J=4.6,1.6Hz,1H),3.88–3.72(m,2H);

[0071] 13 C NMR (125MHz, CDCl3) δ: 171.1, 137.3, 136.4, 128.8, 128.6, 128.4, 128.1, 127.9, 95.7, 88.4, 79.7, 74.7, 74.2, 74.2, 71.7;

[0072] HRMS(ESI)m / z Calcd for C 20 H 21 O6N3Na[M+Na] + 422.1323, found 422.1314.

[0073] Step 3: Preparation and data of compound 4 and compound 5:

[0074] To a 10 mL reaction vial equipped with a magnetic stirrer, compound 3 (500 mg, 1.25 mmol, 1.0 equiv) was added sequentially. Acetone / water solution (v:v = 6:1, 21 mL) was added to dissolve the mixture. Silver carbonate (69 mg, 0.25 mmol, 0.2 equiv), 1-chloromethyl-4-fluoro-1,4-diazabicyclo[2.2.2]octane bis(tetrafluoroborate) salt (selectfluor, 887 mg, 2.5 mmol, 2.0 equiv), and potassium fluoride dihydrate (236 mg, 2.5 mmol, 2.0 equiv) were then added under argon. The mixture was purged three times under argon and reacted at room temperature for 40 min. The mixture was then filtered through a pad of Celite. The filtrate was collected and concentrated under reduced pressure to remove the acetone. The mixture was washed with saturated sodium bicarbonate solution and extracted with EA. The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and the solvent was removed by rotary evaporation. The residue was separated and purified by column chromatography (petroleum ether:ethyl acetate = 2:1) to give colorless syrups 4 and 5 (330 mg, 0.88 mmol, 71%, 1:1.01).

[0075] The analytical data of compound 4 are as follows:

[0076] 1 H NMR(500MHz, CDCl3)δ:7.40–7.27(m,10H),5.39(dd,J=3.7,1.3Hz,1H),4.70–4.58(m,4H ),4.36(dd,J=15.3,4.7Hz,1H),4.07(s,1H),3.86–3.75(m,2H),3.67(t,J=10.3Hz,1H);

[0077] 13 C NMR(125MHz, CDCl3)δ:136.8,136.6,128.7,128.7,128.4,128.4,128.2,128.1,119.8(J C-F =228.6Hz),96.6(J C-F =3.6Hz),81.9(J C-F =32.6Hz),74.3,73.6(J C-F =3.1Hz),73.3,67.7(J C-F =43.9Hz);

[0078] 19 F NMR (470MHz, CDCl3) δ: -106.6 -106.7 (m, 1F);

[0079] HRMS(ESI)m / z Calcd for C19 H 20 O4N3FNa[M+Na] + 396.1330, found 396.1325.

[0080] The analytical data of compound 5 are as follows:

[0081] 1 H NMR(500MHz, CDCl3)δ:7.39–7.28(m,10H),5.59(s,1H),4.71–4.58(m,3H),4.52(d,J=12.0H z,1H),4.29(dd,J=17.1,5.2Hz,1H),3.91(t,J=3.6Hz,1H),3.72–3.60(m,2H),2.78(s,1H);

[0082] 13 C NMR(125MHz, CDCl3)δ:137.5,136.6,128.8,128.7,128.6,128.3,128.1,127.8,116.5(J C-F =233.0Hz),95.8,75.8(J C-F =20.0Hz),73.8,73.4,72.4,68.3(J C-F =43.9Hz);

[0083] 19 F NMR (470MHz, CDCl3) δ: -114.4 (d, J = 16.9Hz, 1F);

[0084] HRMS(ESI)m / z Calcd for C 19 H 20 O4N3FNa[M+Na] + 396.1330, found 396.1319.

[0085] Step 4: Preparation and data of compounds 6a-6d / 7a-7d:

[0086] (1) Preparation and data of compound 6a:

[0087] To a DMF / water mixed solution (v:v = 4:1, 0.25 mL) of compound 4 (10 mg, 0.027 mmol, 1.0 equiv) were added cyclopropylacetylene (6.5 μL, 0.08 mmol, 3.0 equiv), copper sulfate pentahydrate (2.7 mg, 0.011 mmol, 0.4 equiv), and sodium ascorbyl palmitate (4.4 mg, 0.022 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h and then directly loaded onto silica gel for solvent removal. The product was purified by column chromatography (petroleum ether:ethyl acetate = 3:1) to give a colorless syrup 6a (11.2 mg, 0.026 mmol, 95%).

[0088] The analysis data are as follows:

[0089] 1 H NMR(500MHz, CDCl3)δ:7.42(s,1H),7.39–7.22(m,10H),6.10(s,1H),4.93–4.82(m,1H),4.80–4.70(m,2H) ,4.68–4.58(m,2H),4.53(dt,J=14.0,6.8Hz,1H),3.83–3.68(m,3H),2.04–1.83(m,1H),0.99–0.79(m,4H);

[0090] 13 C NMR(125MHz, CDCl3)δ:155.2,136.8,136.8,128.8,128.7,128.5,128.4,128.2,120.4(J C-F =230.8Hz),118.6,94.0,81.2(J C-F =34.8Hz),74.3,74.2,67.9(J C-F =37.1Hz),7.90,7.89,6.78;

[0091] 19 F NMR (470MHz, CDCl3) δ: -106.2 -106.3 (m, 1F);

[0092] HRMS(ESI)m / z Calcd for C 24 H 26 O4N3FNa[M+Na] + 462.1800,found 462.1790.

[0093] (2) Preparation and data of compound 7a:

[0094] To a solution of compound 5 (8 mg, 0.0214 mmol, 1.0 equiv) in DMF / water (v:v = 4:1, 0.25 mL) were added cyclopropylacetylene (5 μL, 0.0642 mmol, 3.0 equiv), copper sulfate pentahydrate (2.1 mg, 0.0086 mmol, 0.4 equiv), and sodium ascorbyl palmitate (3.4 mg, 0.0171 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h and then directly loaded onto silica gel for solvent removal. The product was purified by column chromatography (petroleum ether:ethyl acetate = 3:1) to give a colorless syrup 7a (9.4 mg, 0.0212 mmol, 99%).

[0095] The analysis data are as follows:

[0096] 1 H NMR(500MHz, CDCl3)δ:7.40–7.24(m,9H),7.20(d,J=6.7Hz,2H),6.29(s,1H),4.76–4.62(m,3H),4.62–4.58(m,1H),4.47(d,J=11.6Hz, 1H),4.41(d,J=11.6Hz,1H),3.74–3.64(m,2H),3.07(s,1H),1.84(ddd,J=13.5,8.5,5.1Hz,1H),0.94–0.86(m,2H),0.82–0.70(m,2H);

[0097] 13 C NMR(125MHz, CDCl3)δ:150.6,137.2,136.5,128.9,128.7,128.6,128.4,128.1,127.9,119.4,116.9(J C-F =234.6Hz),93.5,76.3(J C-F =18.0Hz),73.9,73.6,72.5,68.4(J C-F =42.0Hz),7.91,7.90,6.65;

[0098] 19 F NMR (470MHz, CDCl3) δ: -115.2 -115.4 (m, 1F);

[0099] HRMS(ESI)m / z Calcd for C 24 H 26 O4N3FNa[M+Na] +462.1800,found 462.1788.

[0100] (3) Preparation and data of compound 6b:

[0101] To a DMF / water mixed solution (v:v = 4:1, 1.0 mL) of compound 4 (40 mg, 0.11 mmol, 1.0 equiv) were added phenylacetylene (35 μL, 0.32 mmol, 3.0 equiv), copper sulfate pentahydrate (11 mg, 0.043 mmol, 0.4 equiv), and sodium ascorbyl palmitate (17 mg, 0.086 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h. The DMF was then drained, filtered through a pad of silica gel, and purified by column chromatography (petroleum ether:ethyl acetate = 4:1) to give 6b (49 mg, 0.10 mmol, 95%) as a white solid.

[0102] The analysis data are as follows:

[0103] 1 H NMR (400MHz, CDCl3) δ: 7.83 (s, 1H), 7.75-7.68 (m, 2H), 7.38-7.15 (m, 13H), 6.14 (t, J = 4.32, 1H), 4.94-4.83 (m, 1 H),4.73-4.68(m,2H),4.60-4.54(m,2H),4.47(dd,J=9.7,4.8Hz,1H),3.93(d,J=10.1Hz,1H),3.82-3.66(m,2H);

[0104] 13 C NMR(101MHz, CDCl3)δ:148.5,136.9,136.8,130.1,129.0,128.8,128.7,128.6,128.5,128.2,126.0,120.6(J C-F =231.5Hz),118.43,118.41,94.2,81.2,80.9,74.5,74.3,74.2,67.9(J C-F =35.9Hz);

[0105] 19 F NMR (376MHz, CDCl3) δ: -105.9 (s, 1F);

[0106] HRMS(ESI):m / z Calcd for C 27 H 27 FN3O4[M+H] +476.1980,found 476.1966.

[0107] (4) Preparation and data of 7b:

[0108] To a DMF / water mixed solution (v:v = 4:1, 1.0 mL) of compound 5 (40 mg, 0.11 mmol, 1.0 equiv) were added phenylacetylene (35 μL, 0.32 mmol, 3.0 equiv), copper sulfate pentahydrate (11 mg, 0.043 mmol, 0.4 equiv), and sodium ascorbyl palmitate (17 mg, 0.086 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h. The DMF was then drained, filtered through a pad of silica gel, and purified by column chromatography (petroleum ether:ethyl acetate = 4:1) to give 7b (46 mg, 0.11 mmol, 97%) as a white solid.

[0109] The analysis data are as follows:

[0110] 1 H NMR(400MHz, CDCl3)δ:7.82(s,1H),7.59-7.50(m,2H),7.37-7.07(m,15H),6.37(s,1H),4.72 -4.53(m,4H),4.41(d,J=11.5Hz,1H),4.34(d,J=11.5Hz,1H),3.70-3.64(m,2H),3.15(s,1H);

[0111] 13 C NMR(101MHz, CDCl3)δ:148.0,137.1,136.5,130.1,129.0,128.9,128.8,128.7,128.5,128.4,128.2,127.9,125.8,118.9,116.9(J C-F =234.9Hz),93.9,75.9(J C-F =18.1Hz),73.9,73.6,72.6,68.3(J C-F =43.2Hz);

[0112] 19 F NMR (376MHz, CDCl3) δ: -115.4 (s, 1F);

[0113] HRMS(ESI):m / z Calcd for C 27 H 27 FN3O4[M+H] +476.1980,found 476.1968.

[0114] (5) Preparation and data of compound 6c:

[0115] To a DMF / water mixed solution (v:v = 4:1, 1.0 mL) of compound 4 (40 mg, 0.11 mmol, 1.0 equiv) were added 2-methyl-3-butyn-2-ol (31 μL, 0.32 mmol, 3.0 equiv), copper sulfate pentahydrate (11 mg, 0.043 mmol, 0.4 equiv) and sodium ascorbyl palmitate (17 mg, 0.086 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h. The DMF was then drained, filtered through a pad of silica gel, and purified by column chromatography (petroleum ether:ethyl acetate = 1:1) to give 6c (46 mg, 0.10 mmol, 94%) as a white solid.

[0116] The analysis data are as follows:

[0117] 1 H NMR (400MHz, CDCl3) δ: 7.57 (s, 1H), 7.34-7.14 (m, 10H), 6.09 (t, J = 4.5Hz, 1H), 4.83 (s, 1H), 4.70 (s, 2H), 4.58 (d, J = 11.8Hz ,1H),4.54(d,J=11.8Hz,1H),4.43(dd,J=8.9,4.8Hz,1H),4.05-3.91(m,1H),3.80-3.65(m,2H),2.66(s,1H),1.53(s,6H);

[0118] 13 C NMR(101MHz, CDCl3)δ:136.84,136.82,128.74,128.67,128.5,128.3,128.2,128.1,120.4(J C-F =232.0Hz),118.2,93.9,80.9(J C-F =35.2Hz),77.3,74.5,74.2,68.6,68.0,67.8(J C-F =34.9Hz),30.4;

[0119] 19 F NMR (376MHz, CDCl3) δ: -105.7 (s, 1F);

[0120] HRMS(ESI):m / z Calcd for C 24 H29 FN3O5[M+H]+458.2086, found 458.2076.

[0121] (6) Preparation and data of compound 7c:

[0122] To a DMF / water mixed solution (v:v = 4:1, 1.0 mL) of compound 5 (40 mg, 0.11 mmol, 1.0 equiv) were added 2-methyl-3-butyn-2-ol (31 μL, 0.32 mmol, 3.0 equiv), copper sulfate pentahydrate (11 mg, 0.043 mmol, 0.4 equiv) and sodium ascorbyl palmitate (17 mg, 0.086 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h. The DMF was then drained, filtered through a pad of silica gel, and purified by column chromatography (petroleum ether:ethyl acetate = 4:1) to give 7c (44 mg, 0.098 mmol, 89%) as a white solid.

[0123] The analysis data are as follows:

[0124] 1 H NMR(400MHz, CDCl3)δ:7.53(s,1H),7.30–7.20(m,8H),7.14–7.09(m,2H),6.27(s,1H),4.66(d,J=11.6Hz,1H),4.60(d,J=11.6Hz,1H) ,4.56(m,2H),4.39(d,J=11.6Hz,1H),4.33(d,J=11.6Hz,1H),3.62(d,J=5.9Hz,2H),3.23(m,1H),2.47(s,1H),1.44(d,J=7.4Hz,6H);

[0125] 13 C NMR(101MHz, CDCl3)δ:155.8,137.2,136.5,128.8,128.7,128.6,128.4,128.2,127.9,118.9,116.8(J C-F =235.0Hz),93.7,77.4,76.1(J C-F =18.0Hz),73.8,73.5,72.5,68.4,68.3(J C-F =42.5Hz),30.33,30.28;

[0126] 19 F NMR (376MHz, CDCl3) δ: -115.5 (s, 1F);

[0127] HRMS(ESI):m / z Calcd for C 24 H 29 FN3O5[M+H]+458.2086, found 458.2072.

[0128] (7) Preparation and data of 6d:

[0129] To a DMF / water mixed solution (v:v = 4:1, 1.0 mL) of compound 4 (40 mg, 0.11 mmol, 1.0 equiv) were added methyl propiolate (26 μL, 0.32 mmol, 3.0 equiv), copper sulfate pentahydrate (11 mg, 0.043 mmol, 0.4 equiv) and sodium ascorbyl palmitate (17 mg, 0.086 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h, then the DMF was drained, filtered through a pad of silica gel, and purified by column chromatography (petroleum ether:ethyl acetate = 2:1) to give 6d (29 mg, 0.065 mmol, 59%) as a white solid.

[0130] The analysis data are as follows:

[0131] 1 H NMR (400MHz, CDCl3) δ: 8.21 (s, 1H), 7.34-7.17 (m, 10H), 6.15 (t, J = 4.4Hz, 1H), 4.83 (m, 1H) ),4.70(m,2H),4.58(m,2H),4.40(dd,J=9.4,4.8Hz,1H),3.86(s,3H),3.81-3.67(m,3H);

[0132] 13 C NMR(101MHz, CDCl3)δ:160.9,140.6,136.7,136.6,128.8,128.7,128.6,128.4,128.3,128.2,126.5,126.5,120.8(J C-F =232.5Hz),94.4(J C-F =3.5Hz),80.9,80.6,77.4,74.8,74.33,74.30,67.7(J C-F =35.5Hz),52.43;

[0133] 19 F NMR (376MHz, CDCl3) δ: -106.2 (s, 1F);

[0134] HRMS(ESI):m / z Calcd for C 23 H 25 FN3O6[M+H]+458.1722, found 458.1713.

[0135] (8) Preparation and data of 7d

[0136] To a DMF / water mixed solution (v:v = 4:1, 1.0 mL) of compound 5 (40 mg, 0.11 mmol, 1.0 equiv) were added methyl propiolate (26 μL, 0.32 mmol, 3.0 equiv), copper sulfate pentahydrate (11 mg, 0.043 mmol, 0.4 equiv) and sodium ascorbyl palmitate (17 mg, 0.086 mmol, 0.8 equiv) in sequence. The reaction solution was vigorously stirred at room temperature for 1 h, then the DMF was drained, filtered through a pad of silica gel, and purified by column chromatography (petroleum ether:ethyl acetate = 2:1) to give 7d (39 mg, 0.083 mmol, 78%) as a white solid.

[0137] The analysis data are as follows:

[0138] 1 H NMR(400MHz, CDCl3)δ:8.24(s,1H),7.36-7.09(m,10H),6.32(s,1H),4.65-4.57(m,2H),4.57-4.47(m,2H), 4.42(d,J=11.8Hz,1H),4.34(d,J=11.8Hz,1H),3.84(s,3H),3.65-3.56(m,2H),3.08(dd,J=4.2,1.8Hz,1H);

[0139] 13 C NMR (101MHz, CDCl3) δ: 160.8, 140.0, 136.8, 136.3, 128.9, 128.8, 128.6, 128.4, 128.2, 128.0, 127.2, 116.9 (J C-F =235.6Hz),93.8,77.3,75.7(J C-F =19.4Hz),73.8,73.7,72.6,67.8(J C-F =43.0Hz),52.4;

[0140] 19 F NMR (376MHz, CDCl3) δ: -115.7 (s, 1F);

[0141] HRMS(ESI)m / z cald for C 23 H 25 FN3O6[M+H]+458.1722,found 458.1709.

[0142] Example 2: Antiviral activity test

[0143] The prepared 1,2,3-triazole nucleoside compounds were tested for antiviral activity, and it was found that some 1,2,3-triazole nucleosides had good antiviral activity and were excellent antiviral compounds that could be further developed and prepared into antiviral drugs.

[0144] The specific activity tests are as follows:

[0145] 1. Experiment on the inhibition rate of the tested compounds on vesicular stomatitis virus (VSV) at 50 μmol:

[0146] 1.1 Anti-VSV activity test steps:

[0147] (1) Seeding: Vero cells in the logarithmic growth phase were harvested, counted under a microscope, diluted with DMEM complete medium (DMEM + 1% double antibody + 5% fetal bovine serum), and approximately 1.0 × 105 cells were seeded per well in two 24-well plates. The plates were cultured in a 37°C, 5% CO2 incubator for 12 h.

[0148] (2) Virus adsorption: When the virus is diluted to a volume of 200 μL per well using serum-free and double-antibody-free DMEM medium, the VSV-GFP MOI is 0.01. First, the virus is diluted in a gradient manner. 90 μL of culture medium is added to a 1.5 mL centrifuge tube, followed by 10 μL of VSV-GFP (2.209×108 PFU / mL). Gently shake to mix, and repeat twice. Take 81 μL and dilute to 3.6 mL with DMEM. Except for the blank control group, discard the culture medium from each well and add 200 μL of VSV virus. Incubate in a 37°C, 5% CO2 incubator (VSV-GFP is incubated for 0.5 h, and gently shake every 0.5 h to mix.)

[0149] (3) Drug preparation: Dissolve the sample in DMSO to 50 mM. Take 0.6 μL and dilute 1000-fold to 0.6 mL with culture medium without double antibody, with a final concentration of 50 μM. Mix well with vortex. For the positive drug, take 0.3 μL of RBV (40 mM) and dilute it to 0.6 mL with culture medium without double antibody, with a final concentration of 20 μM. For the negative control group, take 0.6 μL of DMSO and dilute it to 0.6 mL with culture medium without double antibody.

[0150] (4) Drug Addition: Aspirate the virus, add 500 μL of DMSO-containing medium to the negative control group, and add 500 μL of drug-containing medium to each well (add one well of drug-containing medium after aspirating the virus from each well). Incubate in a 37°C, 5% CO2 incubator (VSV-GFP culture for 16 h).

[0151] (5) Detect green fluorescence and take photos.

[0152] (6) Use the Guava flow cytometer to collect the percentage of positive cells and mean fluorescence intensity data: add 200 μL PBS to each well for washing and then discard, add 200 μL trypsin to digest the cells, add 400 μL culture medium to stop digestion, and collect them in a 1.5 mL EP tube. Centrifuge at 1300 rpm, 25°C, discard the supernatant, add 100 μL PBS to suspend the precipitate, add 200 μL 10% paraformaldehyde, and let it stand at 4°C for 15 minutes. Centrifuge at 1300 rpm, 25°C, discard the supernatant, add 300 μL PBS to suspend the precipitate, transfer to a 96-well plate, and detect using the Guava flow cytometer.

[0153] 1.2 Experimental results:

[0154] like Figure 1 As shown in the figure, at a concentration of 50 μmol, ribavirin had an inhibition rate of 80% against vesicular stomatitis virus. The inhibition rates of compounds 6a, 7a, 6b, 7b, 6c, 7c, 6d, and 7d against vesicular stomatitis virus were 71%, 83%, 82%, 80%, 82%, 79%, 58%, and 69%, respectively.

[0155] 1.3 Results and Discussion

[0156] At a concentration of 50 μmol, the inhibition rate of this class of compounds was almost comparable to that of the positive control drug ribavirin, demonstrating excellent antiviral activity. Furthermore, given that compound 7a showed the highest activity against vesicular stomatitis virus, further in vitro and in vivo experiments were conducted on it.

[0157] Example 3: Inhibition of RABV proliferation in vitro

[0158] 2.1 Materials and Methods

[0159] 2.1.1 Compounds and Reagents: Compound 7a was dissolved in a 10 mM stock solution containing dimethyl sulfoxide (DMSO) and stored at -20°C.

[0160] 2.1.2 Cells, viruses and antibodies: BSR cells (BHK-21, CCL-10 clones) were cultured in Dulbecco's modified Eagle's medium (DMEM, Gibco, Grand Island, NY) supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, NY). RABV CVS-B2c was serially passaged from the CVS-24 strain and stored in the laboratory. Monoclonal antibodies (mAbs) against RABV nucleoprotein (RABV-n) were generated in our laboratory. A mAb against phosphoprotein (RABV-P) conjugated to FITC (FITC-RABV-P) was generated by Friendbio Technology Co., Ltd. (Wuhan, China). Anti-β-actin mAbs were purchased from ProteinTech (ProteinTech Group, Inc, USA).

[0161] 2.2 Cytotoxicity assay

[0162] The cytotoxicity of compound 7a was analyzed using the MTS (Promega, USA) method according to the manufacturer's instructions. Briefly, BSR cell monolayers were prepared on 96-well plates and then incubated with 100 μL DMEM (2% FBS) containing different concentrations (0, 10, 20, 40, 60, 80, 100, 120 μM) of LM-19 at 37°C for 48 h. After incubation, 20 μL of MTT solution was added and incubated at 37°C for 2 h. The absorbance at 490 nm was then recorded using an automated plate reader (Spectra-Max 190, Molecular Devices, Inc, California, USA).

[0163] 2.3 Viral suppression test

[0164] To evaluate the antiviral effect of compound 7a, BSR cells were cultured overnight in 24-well cell culture dishes. They were infected with RABV at an MOI of 0.01 at 37°C for 1 h. The supernatant was collected and the cells were further cultured with fresh drug-containing medium. At 48 hp / i, the cell supernatant was collected for virus titration and washed with RIPA buffer (Beyotime Biotech Inc., China) and Cells were lysed with ELISA Reagent (Invitrogen Inc., California, USA) for further quantitative analysis.

[0165] 2.4 Virus titration

[0166] Cell supernatants were titrated using a direct fluorescent antibody assay. Briefly, BSR cells in 96-well plates were serially inoculated with 10-fold dilutions of supernatant containing RABV and incubated at 37°C for 48 hours. After incubation, cells were fixed with 80% ice-cold acetone and then stained with FITC-RABV-P antibody. Antigen-positive foci were counted using fluorescence microscopy, and viral titers were calculated as foci units / mL (FFU / mL).

[0167] 2.5 Real-time quantitative PCR (qPCR)

[0168] According to the manufacturer's instructions, use Reagents for the isolation of total RNA from brain tissue and cells. II Q RT SuperMix and II 1st Strand cDNA Synthesis Kit (VazymeBiotech Co., Ltd, China) was used to extract equal amounts of total RNA from each sample to generate cDNA for qPCR detection. qPCR was performed using PCR-PCR-guided PCR with PCR-guided Green Supermix (Bio-Rad, USA). Primers for detecting RABV viral RNA were: forward RABV-vrna-f, 5'-ctccacaacgagatgctcaa-3', reverse 5'-CATCCAACGGGAACAAGACT-3'. Primers for detecting RABV-N mRNA were: forward 5'-GATCGTGGAACACCATACCC-3', reverse 5'-TTCATAAGCGGTGACGACTG-3'. A standard curve was generated using serially diluted plasmid (pcDNA3.1-CVS-B2c), and the copy numbers of RABV-vRNA and N mRNA were normalized to 1 μg of total RNA.

[0169] 2.6 Western blot analysis

[0170] BSR cells were washed with PBS and lysed with 1 mM phenylmethylsulfonyl fluoride (PMSF) in RIPA buffer (Beyotime). Protein concentration was determined using a BCA protein assay kit (Beyotime). Equal amounts of protein lysate were separated on 8% or 10% SDS-polyacrylamide gels and transferred to PVDF membranes (Bio-Rad). The membranes were blocked with 5% (w / v) nonfat dry milk in TBST for 4 hours at room temperature and incubated with primary antibodies diluted in TBST overnight at 4°C. After washing three times with TBST, the membranes were probed with an enzyme-conjugated goat anti-mouse IgG antibody (Boster, China) for 1 hour at room temperature. Signal acquisition and analysis were performed using a Syngene G-Box (Syngene, Frederick, MD). The average density of protein bands was measured using Image J software.

[0171] 2.7 Virus inactivation, adsorption, penetration, replication, and release tests

[0172] Inactivation assay: RABV (MOI = 0.05) was mixed with compound 7a (60 μM) or dimethyl sulfoxide (DMSO) and incubated at 37°C for 3 hours. BSR cells cultured in 24-well plates were pre-chilled at 4°C for 1 hour and incubated at 4°C for 2 hours. Cells were washed three times with cold PBS, overlaid with low-melting agarose mixed with 2x DMEM (1:1), and incubated at 37°C for 48 hours. Fluorescent foci were counted using Image J software after staining with FITC-RABV-P antibody.

[0173] Adsorption assay: BSR cells in 24-well plates were precooled at 4°C for 1 h, and the medium was replaced with a mixture of RABV (MOI = 0.05) and LM-19 (60 μM) or DMSO. After incubation at 4°C for another 2 h, the cells were washed three times with cold PBS and then treated as described in [

[0174] 2.8 Experimental Results

[0175] like Figure 2 As shown in A, compound 7a has a cell viability value of more than 90% at 80 μM. Figure 2 As shown in Figures BE, virus titers and virus-encoded intracellular phosphoprotein (RABV-N) decreased with increasing concentrations of 7a, and the expression of RABV-N and viral RNA (vRNA) was reduced after treatment with compound 7a.

[0176] 2.9 Results and Discussion

[0177] Figure 2 A shows that at a concentration of 80 μM, compound 7a showed no cytotoxicity. 50 The value is as high as 123.6μM. Figure 2As shown in Figures BF, 7a inhibited the proliferation of RABV in a dose-dependent manner. Virus titers and virally encoded intracellular phosphoprotein (RABV-N) decreased with increasing 7a concentrations ( Figure 2 B and C). Western Blotting ( Figure 2 D) and RT-PCR ( Figure 2 E and F) show that the expression of RABV-N and viral RNA (vRNA) was reduced after 7a treatment. In summary, 4'-fluoro-1,2,3-triazole nucleoside analog 7a has strong antiviral activity against RABV.

[0178] Example 4: Plasma pharmacokinetics and tissue distribution studies of compound 7a in mice

[0179] 3.1 Plasma pharmacokinetics of samples injected intramuscularly and intravenously into mice

[0180] Three male BALB / c mice were fasted for 12 hours before the experiment and allowed free access to water. 12.04 mg of the sample to be tested was accurately weighed and prepared into a 3 mg / mL injection solution with 10% DMSO, 40% PEG-300, 5% Tween-80, and 45% PBS buffer. Intramuscular injection (30 mg / kg, 0.1 mL per site × 2 sites) was performed on mice. Blood was collected from the canthal vein using a glass capillary tube (30 μL) and transferred into sodium heparinized EP tubes. Plasma was separated and stored at -40°C for later use.

[0181] Three male BALB / c mice were fasted for 12 hours before the experiment and had free access to water. A 3 mg / mL injection solution was diluted to a concentration of 0.6 mg / mL in 10% DMSO, 40% PEG-300, 5% Tween-80, and 45% PBS buffer. After tail vein injection (3 mg / kg, 5 mL / kg), 30 μL of blood was collected from the canthal vein using a glass capillary tube and transferred into sodium heparinized EP tubes. Plasma was separated and stored at -40°C for later use.

[0182] 3.2 Experimental study on tissue distribution of samples injected intramuscularly into mice

[0183] Twelve male BALB / c mice were fasted for 12 hours before the experiment and allowed free access to water. They were randomly divided into four groups, three mice per group, for a total of four: a blank control group and experimental groups administered 15 minutes, 1 hour, and 4 hours after administration. 9.10 mg of the desired sample was accurately weighed and prepared with 10% DMSO, 40% PEG-300, 5% Tween-80, and 45% PBS buffer to a 3 mg / mL injection concentration. Mice were injected intramuscularly (30 mg / kg, 0.1 mL per site × 2 sites) at 15 minutes, 1 hour, and 4 hours after administration. Mice were anesthetized using a small animal anesthesia machine. Blood was collected from the abdominal aorta or by removing the eyeball into a heparinized EP tube. Plasma was separated by centrifugation at 3000 rpm for 10 minutes. The heart, liver, spleen, lung, kidney, brain, and muscle tissues were removed, washed with saline to remove blood stains, blotted dry, weighed, and homogenized with saline at a 1:3 (w / v) ratio. All samples were stored at -40°C.

[0184] 3.3 Pretreatment of plasma and tissue samples

[0185] Take 10 μL of mouse plasma, add 10 μL of internal standard solution (200 ng / mL) and 90 μL of acetonitrile, vortex mix, centrifuge twice at 14,000 rpm for 10 min, and transfer 40 μL to a liquid chromatography vial for LC-MS analysis. If the plasma drug concentration exceeds the standard curve, dilute the plasma sample 10-fold with blank mouse plasma and process as above.

[0186] Take 10 μL of brain, heart, liver, and kidney tissue homogenate, add 90 μL of the respective blank mouse tissue homogenate, vortex mix, add 100 μL of internal standard solution (200 ng / mL) and 100 μL of acetonitrile, vortex mix, centrifuge twice at 14000 rpm × 10 min, take 80 μL into a liquid phase vial, and perform LC-MS / MS analysis.

[0187] Take 100 μL of spleen, lung and muscle tissue homogenate, add 100 μL of internal standard solution (200 ng / mL) and 100 μL of acetonitrile, vortex mix, centrifuge twice at 14000 rpm × 10 min, take 80 μL into a liquid phase vial for LC-MS / MS analysis.

[0188] 3.4 Experimental Results

[0189] The calculated plasma kinetic parameters are shown in Tables 1 and 2. Figure 3 As shown, after intravenous injection (3 mg / kg) and intramuscular injection (30 mg / kg) of compound 7a were respectively given to mice, the drug in the plasma could be completely cleared after 4-6 hours.

[0190] Figure 4The results showed that compound 7a was distributed in plasma, brain, heart, liver, kidney, spleen, lung, and muscle. 0.25 hours after administration, its highest distribution was in the kidney, followed by the heart, brain, and plasma, and least in the spleen. At 0.25, 1, and 4 hours after administration, tissue and plasma drug concentrations in the kidney and heart were all greater than 1.

[0191] Table 1 Plasma pharmacokinetic parameters of the tested compound 7a (3 mg / kg) after intravenous injection in mice

[0192]

[0193] Table 2 Plasma pharmacokinetic parameters of the tested compound 7a (30 mg / kg) after intramuscular injection in mice

[0194]

[0195] 3.5 Results and Discussion

[0196] Compound 7a exhibits broad and rapid distribution in the body, exhibiting no accumulation in various tissues and capable of penetrating the blood-brain barrier and entering the brain. The ratio of kidney and heart tissue concentration to plasma concentration for compound 7a was greater than 1 at 0.25, 1, and 4 hours after administration, suggesting that the compound accumulates in these tissues and may be primarily excreted by the kidney. As a blood-rich organ, liver tissue concentrations were relatively low during the assay period, suggesting that compound 7a does not significantly bind to liver tissue.

[0197] Although the present invention has been described in detail through the above specific embodiments, it should be understood that the above description is not intended to limit the present invention. After reading the above description, various modifications and substitutions of the present invention will be apparent to those skilled in the art. Therefore, the scope of protection of the present invention should be defined by the appended claims.

Claims

1. A 4'-fluoro-1, 2, 3-triazole nucleoside analogue, characterized in that: The general structural formula is shown below: ； Among them, R 1 and R 2 The substituent is one of benzyl, 4-methoxybenzyl and 4-bromobenzyl; R 3 The substituent is one of cyclopropyl, phenyl, 1-methyl-1-hydroxyethyl, and methoxycarbonyl.

2. The 4'-fluoro-1, 2, 3-triazole riboside analogue according to claim 1, wherein R 1 or R 2 The substituent is benzyl.

3. A method for preparing the 4'-fluoro-1, 2, 3-triazole nucleoside analogue according to claim 1, characterized in that: The following steps are involved: ； R 4 is any one of methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, and tert-butyl; (1) Using the known compound 1 as the starting material, the acetoxy group of 1 was activated under acidic conditions and glycosylated with trimethylsilyl azide to obtain the terminal azide compound 2; (2) Remove R from azide compound 2 under alkaline conditions 4 and acetoxy protecting group to obtain uronic acid 3; (3) Decarboxylation and fluorination of the carboxyl group 3 of uronic acid to obtain compound 4 or compound 5; (4) The azide group of compound 4 or compound 5 is subjected to a "click reaction" with an alkyne to obtain triazole nucleoside analogue 6 or 7.

4. The method for preparing a nucleoside analogue according to claim 2, wherein The glycosidation reaction in step (1) is carried out by reacting compound 1 with trimethylsilyl azide at -78-100°C under the catalysis of proton acid or Lewis acid. o C reaction.

5. The method for preparing a nucleoside analogue according to claim 2, wherein The deprotection group in step (2) is obtained by reacting compound 2 with a mixture of water and an organic solvent at -78-100°C under the action of a base. o C reaction.

6. The method for preparing a nucleoside analogue according to claim 2, wherein: The decarboxylation fluorination in step (3) is carried out in a mixed solvent consisting of water and acetone, wherein compound 3, a catalyst, an additive and a free radical receptor are reacted at 0-100 atm. o Reaction under C conditions.

7. The method for preparing a nucleoside analogue according to claim 2, wherein: The "click reaction" described in step (4) is a reaction of compound 4 or 5 with alkyne, copper salt and sodium ascorbyl palmitate at 0-100 o Reaction under C conditions.

8. Use of the 4'-fluoro-1, 2, 3-triazole riboside analogue according to claim 1 or 2 in the preparation of an anti-rabies virus inhibitor.

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