Preparation and application of carbocyclic nucleoside derivative containing ethylidene

Abstract

The invention relates to the technical field of carbocyclic nucleoside containing an ethyl subunit, a liver targeting prodrug thereof and a pharmaceutical composition containing the nucleoside, and discloses preparation and application of carbocyclic nucleoside derivatives containing the ethyl subunit. Comprising retroviruses such as hepatitis B virus (HBV), human immunodeficiency virus (HIV) and the like and secondary disease states and diseases (liver cirrhosis and liver cancer) of the retroviruses, hepatitis D virus (HDV), herpes simplex virus I and herpes simplex virus II (HSV-I and HSV-2), cytomegalovirus (CMV), varicella-zoster virus (VZV), Epstein-Barr virus (EBV) and secondary cancer caused by the retroviruses. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to carbocyclic nucleosides containing an ethylidene group, their liver-targeting prodrugs, pharmaceutical compositions containing these nucleosides, and their use in the treatment or prevention of various viral infections and their secondary disease states and conditions. Specifically, it relates to the preparation and application of carbocyclic nucleoside derivatives containing an ethylidene group. Background Art

[0002] When humans are infected with hepatitis B virus (HBV), acute or fulminant hepatitis may occur, and in severe cases, death may even occur; chronic hepatitis infection may also occur and progress to cirrhosis, and even ultimately develop into hepatocellular carcinoma in some cases. The estimated number of patients infected with HBV worldwide is about 260 million, and the patients are mainly distributed in Asia, Africa, and some countries in Eastern Europe. China, Southeast Asia, and sub-Saharan Africa are high-prevalence regions of hepatitis B.

[0003] HBV is an incomplete double-stranded DNA virus. In its life cycle, viral DNA replication requires reverse transcription, that is, the synthesis of rcDNA from pgRNA. However, human hepatocytes, which are the hosts of HBV, do not perform reverse transcription. Therefore, by inhibiting reverse transcription at this stage, compounds that can inhibit HBV replication can be found, and then drugs for treating hepatitis B can be developed. Similarly, the human immunodeficiency virus (HIV) that causes acquired immunodeficiency syndrome also replicates through reverse transcription. Many nucleoside derivative preparations that inhibit HBV or HIV replication have been developed by humans.

[0004] Currently, many nucleoside derivative preparations are toxic to host cells and have problems such as side effects or drug resistance due to long-term medication. Therefore, an effective therapy for viral infections such as HBV has not been established. In view of these situations, the present invention aims to provide nucleoside derivatives with antiviral activity and low toxicity to host cells. Summary of the Invention

[0005] (1) Technical Problems to be Solved

[0006] Aiming at the deficiencies of the prior art, the present invention provides a preparation and application of carbocyclic nucleoside derivatives containing an ethylidene group, and solves the problems in the above background art.

[0007] (2) Technical Solutions

[0008] To achieve the above object, the present invention provides the following technical solutions: A preparation and application of carbocyclic nucleoside derivatives containing an ethylidene group. These compounds can exhibit excellent anti-HBV virus activity, and these compounds have no cytotoxicity to the host cells of the HBV virus.

[0009]

[0010] In general formula (I), the base is selected from R is selected from H, methyl, F, Cl, Br, I or R' is selected from H, C1-C6 alkyl, cyclopropyl, benzyl, C2-C18 fatty acyl or benzoyl; R" is selected from H, F, Cl, Br, I, OH, OMe, OEt, OPri, OBn, NH 2 or NHR'.

[0011] R 1 and R 2 are each independently selected from H, C2-C17 fatty acyl, C1-C18 alkyl or ether group, O-linked amino acid residue (D or L), aminophosphate group or R 1 and R 2 together with the oxygen atom to which they are bonded form a carbonic acid diester or phosphoric acid diester group; R 3 is an α-amino acid side chain; R 4 is benzyl, C1-C20 straight-chain alkyl or ether group, C3-C20 branched-chain alkyl or ether group, C3-C6 cycloalkyl or C4-C6 heterocycloalkyl.

[0012] In one embodiment, the compound of formula (I) has the structure of formula (II) and is a carbocyclic nucleoside containing an ethylidene group:

[0013]

[0014] In another embodiment, the compound of formula (I) has the structure of formula (III) and is an aminophosphate of a carbocyclic nucleoside containing an ethylidene group:

[0015]

[0016] wherein R 3 is an α-amino acid side chain, such as the side chains of glycine, alanine, valine, leucine, isoleucine, methionine, proline, tryptophan, serine, tyrosine, cysteine, phenylalanine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine, etc.; R 4 is benzyl, isopropyl, tert-butyl or cyclobutyl ether group.

[0017] In another embodiment, for the carbocyclic nucleoside compound containing an ethylidene group, its salt or the solvate of the compound or the compound salt, the base is selected from the following uracil or thymine analogs:

[0018]

[0019] In another embodiment, the bicyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or the compound salt has a base selected from the following cytosine analogs:

[0020]

[0021] In another embodiment, the bicyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or the compound salt has a base selected from the following adenine analogs:

[0022]

[0023] In another embodiment, the bicyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or the compound salt has a base selected from the following guanine analogs:

[0024]

[0025] In another embodiment, the bicyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or the compound salt is selected from the following bicyclic nucleoside compounds containing an ethylidene group, but not limited to the following compounds:

[0026]

[0027]

[0028]

[0029] Another aspect of the application relates to a pharmaceutical composition comprising a bicyclic nucleoside compound containing an ethylidene group of formula (I), its deuterated form, its salt, or a solvate of the compound or the compound salt.

[0030] Another aspect of the present application relates to a therapeutic agent for treating or preventing viral infections, which is useful in various viral infections and their secondary disease states and disorders, including retroviruses such as human immunodeficiency virus, hepatitis B virus (HBV), human T-cell lymphotropic virus types I, II, and V (HTLV-I, II, V), human foamy virus, and their secondary disease states and disorders (cirrhosis and liver cancer), as well as hepatitis D virus (HDV), herpes simplex virus types I and II (HSV-I and HSV-2), cytomegalovirus (CMV), varicella-zoster virus (VZV), and Epstein-Barr virus (EBV), including drug-resistant viruses and other mutant forms of these viruses.

[0031] Another aspect of the present application relates to a method for treating or preventing hepatitis B. The method comprises administering to a subject in need of treatment a therapeutically effective amount of a compound of formula (I)-(III), its deuterated form, its salt, or a solvate of the compound or the compound salt.

[0032] (III) Beneficial effects

[0033] Compared with the prior art, the present invention provides the preparation and application of a carbocyclic nucleoside derivative containing an ethylidene group, and has the following beneficial effects:

[0034] For the preparation and application of the carbocyclic nucleoside derivative containing an ethylidene group, the carbocyclic nucleoside derivative containing an ethylidene group has a good ability to inhibit the replication of hepatitis B virus. The in vitro antiviral activity of some compounds is superior to that of the clinically used drug lamivudine, and has further development prospects. Detailed implementation manners

[0035] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0036] For the preparation and application of a carbocyclic nucleoside derivative containing an ethylidene group, the carbocyclic nucleoside derivative containing an ethylidene group can exhibit excellent anti-HBV virus activity, and this type of compound has no cytotoxicity to the host cells of HBV virus.

[0037]

[0038] In general formula (I), the base is selected from R is selected from H, methyl, F, Cl, Br, I or R' is selected from H, C1-C6 alkyl, cyclopropyl, benzyl, C2-C18 fatty acyl or benzoyl; R" is selected from H, F, Cl, Br, I, OH, OMe, OEt, OPri, OBn, NH 2 or NHR'.

[0039] R 1 、R 2 are each independently selected from H, C2-C17 fatty acyl, C1-C18 alkyl or ether group, O-linked amino acid residue (D or L), aminophosphate group or R 1 and R 2 together with the oxygen atom to which they are bonded form a carbonic acid diester or phosphoric acid diester group; R 3is an α-amino acid side chain; R 4 is benzyl, a C1-C20 straight-chain alkyl or ether group, a C3-C20 branched alkyl or ether group, a C3-C6 cycloalkyl group, or a C4-C6 heterocycloalkyl group.

[0040] In one embodiment, the compound of formula (I) has the structure of formula (II) and is a carbocyclic nucleoside containing an ethylidene group:

[0041]

[0042] In another embodiment, the compound of formula (I) has the structure of formula (III) and is an aminophosphate of a carbocyclic nucleoside containing an ethylidene group:

[0043]

[0044] wherein R 3 is an α-amino acid side chain, such as the side chains of glycine, alanine, valine, leucine, isoleucine, methionine, proline, tryptophan, serine, tyrosine, cysteine, phenylalanine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine, and histidine, etc.; R 4 is benzyl, isopropyl, tert-butyl, or cyclobutyl ether group.

[0045] In another embodiment, for the carbocyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or compound salt, the base is selected from the following uracil or thymine analogs:

[0046]

[0047] In another embodiment, for the carbocyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or compound salt, the base is selected from the following cytosine analogs:

[0048]

[0049] In another embodiment, for the carbocyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or compound salt, the base is selected from the following adenine analogs:

[0050]

[0051] In another embodiment, for the carbocyclic nucleoside compound containing an ethylidene group, its salt, or a solvate of the compound or compound salt, the base is selected from the following guanine analogs:

[0052]

[0053] In another embodiment, the ethylidene-containing carbocyclic nucleoside compound, its salt, or a solvate of the compound or compound salt is selected from the following ethylidene-containing carbocyclic nucleoside compounds and is not limited to the following compounds:

[0054]

[0055]

[0056]

[0057] Another aspect of the application relates to a pharmaceutical composition, which comprises an ethylidene-containing carbocyclic nucleoside compound of formula (I), its deuterated form, its salt, or a solvate of the compound or compound salt.

[0058] Another aspect of the present application relates to a therapeutic agent for treating or preventing viral infections, which is useful in a variety of viral infections and their secondary disease states and conditions, including retroviruses such as human immunodeficiency virus, hepatitis B virus (HBV), human T-lymphotropic virus types I, II, and V (HTLV-I, II, V), human foamy virus, and their secondary disease states and conditions (cirrhosis and liver cancer), as well as hepatitis D virus (HDV), herpes simplex virus types I and II (HSV-I and HSV-2), cytomegalovirus (CMV), varicella-zoster virus (VZV), and Epstein-Barr virus (EBV), including drug-resistant viruses and other mutant forms of these viruses.

[0059] Another aspect of the present application relates to a method for treating or preventing hepatitis B. The method comprises administering to a subject in need of treatment a therapeutically effective amount of a compound of formula (I)-(III), its deuterated form, its salt, or a solvate of the compound or compound salt.

[0060] Example 1

[0061]

[0062] Intermediate 1

[0063] Add D-(+)-ribonolactone (20.0 g, 0.151 mol) and dry pyridine (200 mL) to a 500 mL round-bottom flask. Slowly add 1,3-dichloro-1,1,3,3-tetraisopropyldisiloxane (53.2 mL, 0.166 mol) dropwise at -10 °C. After warming to 25 °C, stir overnight. After TLC detection shows the reaction is complete, add methanol (20 mL) to terminate the reaction. Evaporate the solvent under reduced pressure. Dissolve the residue in ethyl acetate (300 mL). Wash the organic phase with 5% aqueous citric acid solution and saturated brine successively. Dry over anhydrous sodium sulfate, filter, and concentrate. Purify the residue by silica gel column chromatography (ethyl acetate: petroleum ether = 1:20) to obtain a colorless oily product 1 (47.6 g, 85%). 1H NMR (400 MHz, Chloroform-d) δ 4.66 - 4.60 (m, 1H), 4.23 - 4.19 (m, 1H), 4.14 (dd, J = 12.3, 3.6 Hz, 1H), 3.93 (dd, J = 12.3, 6.6 Hz, 1H), 2.86 (dd, J = 17.3, 8.0 Hz, 1H), 2.71 (dd, J = 17.3, 9.2 Hz, 1H), 1.06 (m, 28H).

[0064] Intermediate 2b

[0065] Dissolve compound 1 (16.0 g, 42.7 mmol) in dry tetrahydrofuran (200 mL). Cool to -78 °C and slowly add diisobutylaluminum hydride (1.5 M, 31.3 mL, 46.9 mmol) dropwise. After the addition, stir at low temperature for 1.5 hours. After TLC monitoring shows the reaction is complete, slowly add propargylmagnesium bromide Grignard reagent (0.5 M, 213 mL, 107 mmol). Let it warm to 25 °C naturally and stir overnight. After TLC detection shows the reaction is complete, slowly add saturated potassium sodium tartrate (200 mL) to terminate the reaction. Stir the mixture vigorously overnight, filter under reduced pressure through a pad of diatomaceous earth, and wash with ethyl acetate (50 mL x 3). Extract the separated aqueous phase with ethyl acetate (100 mL x 2). Combine the organic phases, wash with saturated brine, dry over anhydrous sodium sulfate, filter, and concentrate. Purify the residue by silica gel column chromatography (ethyl acetate: petroleum ether = 1:3) to obtain a light yellow oily substance 2b (9.92 g, 56%). 11H NMR (400 MHz, Chloroform-d) δ 4.58 (d, J = 6.3 Hz, 1H), 4.19 (d, J = 11.6 Hz, 1H), 4.10 (ddd, J = 9.2, 6.8, 4.4 Hz, 2H), 3.80 (dd, J = 11.7, 2.2 Hz, 1H), 3.48 (t, J = 8.5 Hz, 1H), 2.74 (d, J = 9.4 Hz, 1H), 2.18 - 2.06 (m, 2H), 1.79 (d, J = 2.1 Hz, 3H), 1.13 - 0.96 (m, 28H); 13 13C NMR (101 MHz, Chloroform-d) δ 81.51, 80.20, 75.52, 68.04, 62.39, 59.99, 44.28, 17.75, 17.49, 17.41, 17.35, 13.50, 13.36, 12.68, 12.55, 3.64.

[0066] Intermediate 3

[0067] Dissolve compound 2b (3.78 g, 9.07 mmol), EDC·HCl (2.09 g, 10.9 mmol), benzoic acid (1.22 g, 10.0 mmol) and DMAP (111 mg, 0.91 mmol) in 40 mL of dichloromethane. Slowly add triethylamine (1.64 mL, 11.8 mmol) dropwise at -20 °C. After addition, stir the reaction at room temperature for 16 hours. Quench the reaction with water. Extract the aqueous phase twice with dichloromethane (20 mL x 2), dry over anhydrous sodium sulfate, filter and concentrate. After drying the residue, dissolve it in 20 mL of dichloromethane. Add DMAP (94 mg, 0.768 mmol), and add thiocarbonyl diimidazole (2.73 g, 15.4 mmol) under an ice-water bath. Stir the reaction solution at room temperature for 12 hours. Monitor the reaction by TLC until completion. After concentration, separate the residue by silica gel column chromatography (ethyl acetate:petroleum ether = 1:3) to obtain the product 3 as a yellowish-white solid (4.53 g, 82%). 1 1H NMR (400 MHz, Chloroform-d) δ 8.35 (s, 1H), 8.02 (d, J = 7.7 Hz, 2H), 7.67 (s, 1H), 7.55 (t, J = 7.4 Hz, 1H), 7.41 (t, J = 7.7 Hz, 2H), 7.07 (s, 1H), 5.79 (t, J = 7.7 Hz, 1H), 5.38 (d, J = 9.6 Hz, 1H), 4.56 (dq, J = 9.9, 4.9, 3.9 Hz, 1H), 4.19 (s, 2H), 2.16 (q, J = 5.2, 3.7 Hz, 2H), 1.84 (s, 3H), 1.21 - 1.01 (m, 28H);13 C NMR (101 MHz, Chloroform-d) δ 183.06, 165.54, 136.80, 133.30, 131.09, 129.96, 129.85, 128.45, 118.31, 85.09, 83.21, 64.66, 62.24, 58.94, 40.09, 17.90, 17.48, 17.45, 17.31, 17.29, 17.27, 17.21, 13.77, 13.37, 12.78, 12.51, 3.78.

[0068] Intermediate 4

[0069] Under nitrogen protection, a toluene solution of compound 3 (1.31 g, 3.10 mmol), tributyltin hydride (1.00 mL, 3.72 mmol) and AIBN (254 mg, 1.55 mmol) was bubbled to remove oxygen for 20 minutes, and then reacted at 110 °C for 6 hours. The reaction was monitored by TLC until completion. After concentration under reduced pressure, the residue was separated by silica gel column chromatography (ethyl acetate: petroleum ether = 1:20) to obtain a colorless oily product 4 (850 mg, 81%). 1 H NMR (400 MHz, Chloroform-d) δ 7.94 - 7.89 (m, 2H), 7.44 - 7.38 (m, 1H), 7.32 - 7.26 (m, 2H), 5.37 - 5.49 (m, 1H), 5.40 (t, J = 7.2 Hz, 1H), 5.42 - 5.38 (m, 1H), 3.97 - 3.93 (m, 1H), 3.92 - 3.81 (m, 1H), 2.64 - 2.46 (m, 1H), 1.72 - 1.57 (m, 2H), 1.56 - 1.49 (m, 3H), 0.97 - 0.87 (m, 28H).

[0070] Intermediate 5

[0071] Compound 4 (500 mg, 0.99 mmol) was dissolved in anhydrous methanol (5.0 mL), potassium carbonate (273 mg, 1.98 mmol) was added, and the mixture was stirred at room temperature for 5 hours. The solvent was removed by evaporation under reduced pressure, and the residue was separated by silica gel column chromatography (ethyl acetate: petroleum ether = 1:4) to obtain a colorless oily product 5 (350 mg, 88%). Two-dimensional NMR detection of the obtained product found that:

[0072] 5a (minor component, E-form double bond) 11H NMR (400 MHz, Chloroform-d) δ 5.57 (d, J = 7.4 Hz, 1H), 4.32 - 4.29 (m, 1H), 4.05 - 4.03 (m, 1H), 3.48 - 3.45 (m, 1H), 2.88 (d, J = 10.9 Hz, 1H), 2.47 - 2.41 (m, 1H), 1.67 (d, J = 7.8 Hz, 3H), 1.57 - 1.54 (m, 1H), 1.09 - 1.07 (s, 28H).

[0073] 5b (major component, Z-form double bond) 1 1H NMR (400 MHz, Chloroform-d) δ 5.47 (d, J = 7.4 Hz, 1H), 4.66 (d, J = 6.6 Hz, 1H), 4.12 (q, J = 7.2 Hz, 1H), 4.04 - 3.98 (m, 1H), 3.75 (dd, J = 12.1, 6.0 Hz, 1H), 2.63 (d, J = 8.7 Hz, 1H), 2.33 - 2.27 (m, 1H), 1.81 - 1.78 (m, 1H), 1.77 (d, J = 7.0 Hz, 3H), 1.03 - 1.01 (m, 28H).

[0074] Intermediate 7

[0075] At 0 °C, diisopropyl azodicarboxylate (120 μL, 0.60 mmol) was slowly added dropwise to a solution of triphenylphosphine (158 mg, 0.60 mmol), compound 5 (120 mg, 0.30 mmol), and N,N - bis - Boc - protected 6 - chloro - 2 - aminopurine 6 (166 mg, 0.45 mmol) in THF (3.0 mL). After removing the ice bath, the reaction mixture was stirred at room temperature overnight. The solvent was removed under reduced pressure, and the residue was separated by silica gel column chromatography (ethyl acetate:petroleum ether = 1:3) to give the colorless oily product 7 (167 mg, 75%); MS m / e: 751 (M + H) + .

[0076] Intermediates 8a and 8b

[0077] Compound 7 (80 mg, 0.106 mmol) was dissolved in 50% aqueous formic acid (5.0 mL) and stirred at 80 °C for 5 hours. After cooling, the solvent was removed under reduced pressure, and the residue was added to saturated ammonia - methanol solution (5.0 mL) and stirred at room temperature overnight. After concentrating the reaction mixture, AD-RH (amylose tris-3,5-dimethyl phenyl-carbamate; 150 x 10 mm, particle size 5 μm) reverse-phase chiral column, gradient elution (MeCN / H2O: 20%-60%, 5 mL / min) was used for purification by chiral column and reverse-phase chromatography mode to obtain white solids 8a (2 mg, 6%) and 8b (25 mg, 80%).

[0078] 8a: 1 H NMR (400 MHz, DMSO-d 6 ) δ 7.50 (s, 1H), 5.65 (d, J = 7.3 Hz, 1H), 5.15 - 5.02 (m, 1H), 4.22 - 4.25 (m, 1H), 3.48 (qd, J = 11.0, 5.2 Hz, 2H), 2.38 (s, 1H), 2.01 - 2.10 (m, 1H), 1.89 - 1.97 (m, 1H), 1.30 (s, 3H); HRMS (ESI) for C 12 H 18 N 3 O 3 (M + H) + Calculated value 252.1348, found value 252.1333.

[0079] 8b: 1 H NMR (400 MHz, DMSO-d 6 ) δ 7.64 (s, 1H), 5.68 (d, J = 7.3 Hz, 1H), 5.35 - 5.25 (m, 1H), 4.12 (q, J = 5.1 Hz, 1H), 3.63 (qd, J = 11.0, 5.2 Hz, 2H), 2.40 (s, 1H), 2.01 (dq, J = 26.4, 7.6, 7.0 Hz, 2H), 1.17 (d, J = 7.0 Hz, 3H); 13 C NMR (101 MHz, DMSO-d 6 ) δ 156.81, 153.47, 150.76, 140.23, 135.90, 122.98, 116.10, 70.38, 61.67, 54.72, 52.28, 41.60, 13.39.

[0080] Using the same method, intermediate 5 was coupled with cytosine base, thymine base and adenine base respectively to obtain the following compounds 9a, 9b, 10 and 11:

[0081] 1 H NMR (400 MHz, Methanol-d 4)δ8.04(br,1H),7.64(d,J=6.8Hz,1H),5.83(d,J=7.4Hz,2H),5.37(s,1H),4.36 - 4.38(m,1H),3.61 - 3.67(m,2H),2.83 - 2.85(m,1H),2.06 - 2.12(m,1H),1.89 - 1.96(m,1H),1.30(d,J=6.9Hz,3H); HRMS(ESI) for C 13 H 18 N 5 O 3 (M + H) + Calculated 292.1410, found 292.1413.

[0082] 1 H NMR(400MHz, Methanol - d 4 )δ8.39(br,1H),7.79(d,J=6.8Hz,1H),5.83(d,J=7.4Hz,2H),5.62 - 5.59(m,1H),4.25 - 4.22(m,1H),3.80 - 3.78(m,2H),2.49(s,1H),2.23 - 2.16(m,1H),2.01 - 1.96(m,1H),1.46(d,J=6.9Hz,3H); 13 C NMR(101MHz, Methanol - d4)δ165.69,157.24,143.59,139.19,124.52,94.50,71.24,61.73,56.57,54.76,40.63,12.97; HRMS(ESI) for C 13 H 18 N 5 O 3 (M +

[0083] H) + Calculated 292.1410, found 292.1400.

[0084] 1 H NMR(400MHz, Methanol - d 4)δ8.29(d, J = 2.9 Hz, 1H), 8.23(d, J = 2.8 Hz, 1H), 5.93 - 5.82(m, 1H), 5.69(d, J = 7.0 Hz, 1H), 4.39 - 4.30(m, 1H), 4.01 - 3.83(m, 2H), 2.59(s, 1H), 2.32 - 2.24(m, 2H), 1.29(d, J = 6.9 Hz, 3H); 13 C NMR(101 MHz, Methanol - d 4 )δ156.54, 152.20, 149.92, 142.33, 139.82, 126.49, 120.01, 72.75, 63.09, 55.83, 55.31, 42.63, 14.02; HRMS(ESI) for C 13 H 18 N 5 O 2 (M + H) + Calculated value: 276.1460, Measured value: 276.1461.

[0085] 1 H NMR(400 MHz, Methanol - d 4 )δ7.64(s, 1H), 5.85 - 5.73(m, 1H), 5.54 - 5.50(m, 1H), 4.25 - 4.22(m, 1H), 3.79(s, 2H), 2.44(s, 1H), 2.11 - 2.00(m, 2H), 1.79(s, 3H), 1.45(d, J = 7.0 Hz, 3H); 13 C NMR(101 MHz, Methanol - d 4 )δ

[0086] 166.43, 152.90, 140.35, 140.25, 125.80, 111.38, 72.71, 63.24, 56.70, 55.99, 41.51, 14.17, 12.44; HRMS(ESI) for C 13 H 19 N 2 O 4 (M + H) + Calculated value: 267.1345, Measured value: 267.1342.

[0087] Example 2

[0088]

[0089] To a 10 mL single-necked flask, add nucleoside 9b (58 mg, 0.20 mmol) and 1.0 mL of anhydrous THF. The mixture is cooled to 0 °C in an ice-water bath. Dropwise add tert-butylmagnesium chloride Grignard reagent (1 M, 0.6 mL, 0.6 mmol). The reaction mixture is stirred at 0 °C for 30 min, and then a solution of phosphorus reagent 12 (145 mg, 0.32 mmol) in 1 mL of THF is added dropwise. The resulting clear reaction solution is warmed to 35 °C and stirred for 1 day. Add saturated NH4Cl (3 mL) and stir for 5 minutes. Dilute the mixture with ethyl acetate (60 mL). Separate the organic phase, and extract the aqueous layer with ethyl acetate (10 mL). The combined organic layers are washed with water (10 mL), saturated NaHCO3 (2 x 10 mL), brine (10 mL), and dried over Na2SO4. Evaporate the solvent under reduced pressure, and purify the residue by silica gel column chromatography (dichloromethane with 0 - 5% methanol) to obtain the white solid product 13 (42 mg, 41%); 1 H NMR (400 MHz, CDCl 3 ) δ 9.85 (s, 1H), 8.36 (br, 1H), 7.75 (d, J = 6.8 Hz, 1H), 7.15 - 7.34 (m, 5H), 5.83 (d, J = 7.2 Hz, 1H), 5.62 - 5.59 (m, 1H), 4.25 - 4.22 (m, 3H), 3.80 - 3.78 (m, 2H), 2.49 (s, 1H), 2.23 - 2.16 (m, 1H), 2.01 - 1.96 (m, 1H), 1.46 (d, J = 6.9 Hz, 3H), 1.37 (d, J = 7.0 Hz, 3H), 1.23 (d, J = 6.2 Hz, 6H); HRMS (ESI) for C 24 H 34 N 4 O 7 P (M + H) + Theoretical value 521.2087, found value 521.2091.

[0090] Example 3 Fluorescent Quantitative PCR for Detection of HBV DNA

[0091] According to 6.0×10 4Cells / Wells: Seed HepG2.2.15 cells into a 24-well culture plate and continue to culture overnight in an incubator until they adhere and grow. Aspirate the culture medium, and add DMEM medium containing drugs at final concentrations of 0, 0.0064, 0.032, 0.16, 0.8, 4, and 20 μM, 600 μl per well, with 3 replicates for each treatment. Replace the fresh medium containing the same concentration of drugs every 3 days, and extract HBV DNA from the collected medium each time using a hepatitis B virus nucleic acid extraction kit, and store it at -20°C for later use. After 6 days of drug treatment, collect the culture medium supernatant and cells respectively, extract HBV DNA using a hepatitis B virus nucleic acid extraction kit, and detect the copy number of HBV DNA at each concentration by QPCR method, and then calculate the inhibition rate and EC 50 .

[0092] HBV DNA inhibition rate = (copies in cell control group - copies in drug treatment group) / copies in cell control group × 100%

[0093] The EC 50 values of each drug are as follows:

[0094] Compound <![CDATA[EC 50 (μM)]]> Compound <![CDATA[EC 50 (μM)]]> 8a 0.002 8b 0.08 9a 0.07 9b 0.1 10 4.2 11 >20 13 0.005 Lamivudine 0.12

[0095] The experimental results confirm that, compared with the positive control lamivudine, the antiviral activities of compounds 8 and 9 are better. In addition, after the nucleoside compound 10 is converted into its corresponding amino phosphonate prodrug 13, the anti-hepatitis B virus activity is greatly improved, indicating that nucleoside 10 is insensitive to mononucleotide kinase, which affects its ability to be further activated into triphosphate. In summary, these carbocyclic nucleoside derivatives containing ethylidene invented by us have good ability to inhibit hepatitis B virus replication, and the in vitro antiviral activities of some compounds are better than the clinically used lamivudine, showing further development prospects.

[0096] Although the embodiments of the present invention have been shown and described, it will be understood by those of ordinary skill in the art that various changes, modifications, substitutions, and variations can be made to these embodiments without departing from the principles and spirit of the present invention, and the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A preparation and application of a carbocyclic nucleoside derivative containing an ethylene group, characterized in that: A carbocyclic nucleoside derivative containing an ethylene group as shown in general formula (I), a deuterated product thereof, a salt thereof, a solvate of the compound or the salt of the compound: Among them, the base is selected from R is selected from H, methyl, F, Cl, Br, I or R' is selected from H, C1-C6 alkyl, cyclopropyl, benzyl, C2-C18 fatty acyl or benzoyl; R" is selected from H, F, Cl, Br, I, OH, OMe, OEt, OPri, OBn, NH2 or NHR'; R1 and R2 are each independently selected from H, C2-C18 fatty acyl, C1-C20 alkyl or ether group, O-linked amino acid residue (D or L), aminophosphoryl ester group Or R1 and R2 and the oxygen atom to which they are bonded together form a carbonic acid diester or a phosphodiester group; R3 is an α-amino acid side chain; R4 is a benzyl group, a C1-C20 straight-chain alkyl group or an ether group, a C3-C20 branched alkyl group or an ether group, a C3-C6 cycloalkyl group or a C4-C6 heterocycloalkyl group.

2. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claim 1, characterized in that: The compound, its deuterated product, its salt, the solvate of the compound or the salt of the compound, wherein R1 and R2 are H, has a structure shown in formula (II): The definition of base is as described in claim 1.

3. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claim 1, characterized in that: The compound, its deuterated product, its salt, the compound or the solvate of the compound salt has a structure shown in formula (III): Wherein R3 is an α-amino acid side chain, such as glycine, alanine, valine, leucine, isoleucine, methionine (methionine), proline, tryptophan, serine, tyrosine, cysteine, phenylalanine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine and histidine; R4 is benzyl, isopropyl, tert-butyl or cyclobutyl ether; the definition of the base is as described in claim 1.

4. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claim 1, characterized in that: The compound, its deuterated product, its salt, the solvate of the compound or the salt of the compound, the base is selected from the following uracil or thymine analogs:

5. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claim 1, characterized in that: The compound, its deuterated substance, its salt, the compound or the solvate of the compound salt, the base is selected from the following cytosine analogs:

6. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claim 1, characterized in that: The compound, its deuterated product, its salt, the solvate of the compound or the salt of the compound, the base is selected from the following adenine analogs:

7. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claim 1, characterized in that: The compound, its deuterated product, its salt, the solvate of the compound or the salt of the compound, the base is selected from the following guanine analogs:

8. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claims 1-7, characterized in that: The carbocyclic nucleoside compounds containing ethylene groups are selected from the following compounds but are not limited to the following compounds:

9. The preparation and application of a carbocyclic nucleoside derivative containing an ethylene group according to claims 1-8, characterized in that: The compound, its deuterated substance, its salt, or a solvate of the compound or its salt is used as an active ingredient.

10. The preparation and use of a carbocyclic nucleoside derivative containing an ethylene group according to claim 9, characterized in that: The compounds, their deuterated substances, their salts, solvates of the compounds or their salts, and pharmaceutical compositions containing these nucleosides are useful in treating or preventing a variety of viral infections and their secondary disease states and conditions, including human immunodeficiency virus (HIV), hepatitis B virus (HBV), human T-cell lymphotropic virus types I, II and V (HTLV-I, II, V), retroviruses such as human foamy virus and their secondary disease states and conditions (liver cirrhosis and liver cancer), as well as hepatitis delta virus (HDV), herpes simplex virus I and II (HSV-I and HSV-2), cytomegalovirus (CMV), varicella-zoster virus (VZV) and Epstein-Barr virus (EBV), including drug-resistant viruses and other mutant forms of these viruses.

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