Novel anti-cancer drug TNBG-sugar conjugate as well as preparation and application thereof

Abstract

The invention provides a novel anti-cancer drug TNBG-sugar conjugate as well as preparation and application thereof, and belongs to the field of chemical medicines. The conjugates are compounds as shown in a formula I and a formula IV, or salts or stereoisomers thereof. The anticancer drug TNBG-sugar conjugate and a preparation prepared from the anticancer drug TNBG-sugar conjugate matched with appropriate auxiliary materials or auxiliary components have a good inhibition effect on cells of cancers such as lung cancer and liver cancer, are low in toxicity or basically non-toxic to normal cells, can be used for preventing and treating the cancers such as the lung cancer and the liver cancer, and have a good application prospect.

Description

Technical Field

[0001] The invention belongs to the field of chemical medicine and relates to a novel anticancer drug TNBG-sugar conjugate and its preparation and application. Background Art

[0002] Sugar, also known as carbohydrates, has the advantages of low toxicity and good biocompatibility, especially playing an important role in biological processes such as cell recognition, microbial infection, immune response and cell proliferation. As a novel treatment method, drug-sugar conjugates show high efficiency and specificity. At present, more than 170 drug-sugar conjugates have been successfully approved by the Food and Drug Administration of the United States and other countries for marketing, including sugars (oligosaccharides / polysaccharides), small molecule glycosides and sugar mimetics, glycopeptides and glycoproteins, and sugar-based vaccines. Existing studies have shown that in anti-tumor treatment, anticancer drug-sugar conjugates can play their anti-cancer effects, and their effects are similar to the combination of drugs and antibodies. This combination method can increase the solubility of the drug, its stability in the body, prolong the duration of the drug's action, and improve the therapeutic effect by modifying sugars; in addition, sugars can be used as targeting molecules to recognize and bind to tumor cells through specific receptors or sugar chain structures on the cell surface, thereby achieving the purpose of treatment. This not only has high specificity, but also reduces damage to normal cells and improves the safety and effectiveness of treatment. In short, as drug-sugar conjugates are used more and more widely in the medical field, we firmly believe that they will make greater contributions to human health in the future.

[0003] Tetrazanbigen (TNBG) is a new type of azasteroid compound with good anti-tumor activity in vitro and in vivo, and has no cross-resistance with other commonly used anti-tumor drugs. Experimental studies have shown that TNBG can activate the transcription factor SREBP1 (Sterolregulated element-binding protein transfer protein) and peroxisome proliferator-activated receptor γ (PPARγ), inhibit lipid transport MTTP (Microsomal triglyceridtransferprotein), cause lipid aggregation in tumor cells, and tumor cells die due to "lipotoxicity". Therefore, TNBG is a new type of non-cytotoxic anti-tumor drug that causes lipid aggregation in tumor cells with multi-target specificity to produce "lipotoxic" anti-tumor. However, due to the strong lipid solubility of TNBG and poor water solubility, it affects absorption, blood drug concentration and tissue concentration, which seriously limits its drugability.

[0004] In summary, the strategy of conjugating the anticancer drug TNBG with sugars can improve the water solubility of TNBG and enhance its antitumor activity. To this end, the design and development of new anticancer drug TNBG-sugar conjugates will surely have a very important role and significance. Summary of the invention

[0005] The present invention is based on the anticancer drug TNBG, and through the modification of its structure, a series of new anticancer drug TNBG-sugar conjugates with good drugability are innovatively obtained.

[0006] One object of the present invention is to provide a preparation method and use of the novel anticancer drug TNBG-sugar conjugates shown in Formula I and Formula IV.

[0007] Another object of the present invention is to provide a method for producing the compounds of Formula I and Formula IV, or salts thereof, or stereoisomers thereof.

[0008] To achieve the above purpose, the present invention provides the following technical solutions:

[0009] In the first aspect, the present invention provides an anticancer drug TNBG-sugar conjugate or a pharmaceutically acceptable salt or a stereoisomer thereof as shown in Formula I:

[0010]

[0011] in,

[0012] n is an integer selected from 1 to 6;

[0013] G is selected from monosaccharides (glucose, galactose, mannose, etc.), polysaccharides (dextran, lactose, etc.), etc.;

[0014] R1-R8 is selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxyl, nitro, amino, carboxyl, formula II or formula III, etc.;

[0015]

[0016] n1 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond;

[0017] R9 and R 10 Selected from hydrogen, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl; and methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl having a substituent;

[0018] n2 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond;

[0019] n3 is an integer selected from 1 to 2, the symbol represents a single bond or a double bond;

[0020] X is selected from oxygen, nitrogen or carbon;

[0021] R 11 Selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxyl, nitro, amino or carboxyl, etc.

[0022] In some preferred embodiments, the compound of formula I or its pharmaceutically acceptable salt or stereoisomer thereof, wherein n is often selected from the integer 3, and G can be selected from D-glucose, D-galactose, etc.

[0023] The present invention provides a synthetic route for the anticancer drug TNBG-sugar conjugate shown in Formula I, as follows:

[0024]

[0025] The synthesis of the anticancer drug TNBG-sugar conjugate shown in Formula I has the following characteristics:

[0026] First, the synthesis was performed according to the mature route of the research group to obtain the lead compound of the anticancer drug TNBG, i.e., the key intermediate 10, and then the target compound of formula I was obtained through substitution reaction, glycosylation reaction, and deacetylation reaction;

[0027] The substitution reaction is that the hydrogen on the secondary amine is replaced by 1-bromoethanol or propanol under strong alkaline conditions (NaH (60%)); the glycosidation reaction is that the hydroxyl group reacts with the chirally fixed tetraacetyl brominated sugar under the catalysis of heavy metal salts; the deacetylation reaction is carried out in a methanol solution in an alkaline (potassium carbonate) environment to obtain the target compounds I-1 and I-2.

[0028] In a second aspect, the present invention provides an anticancer drug TNBG-sugar conjugate or a pharmaceutically acceptable salt or a stereoisomer thereof as shown in Formula IV:

[0029]

[0030] in,

[0031] n is an integer selected from 1 to 6;

[0032] G is selected from monosaccharides (glucose, galactose, mannose, etc.), polysaccharides (dextran, lactose, etc.), etc.;

[0033] R1-R4 is selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxyl, nitro, amino, carboxyl, formula V or formula VI, etc.;

[0034]

[0035] n1 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond;

[0036] R5 and R6 are selected from hydrogen, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl; and methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl having a substituent;

[0037] n2 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond;

[0038] n3 is an integer selected from 1 to 2, the symbol represents a single bond or a double bond;

[0039] X is selected from oxygen, nitrogen or carbon;

[0040] R7 is selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxy, nitro, amino or carboxyl, etc.

[0041] In some preferred embodiments, the compound of formula IV or its pharmaceutically acceptable salt or stereoisomer thereof, wherein n is often selected from integers 2 and 3; R 1-4 wherein is hydrogen, alkyl is preferably methyl, halogen is preferably F, Cl, alkoxy is preferably methoxy, etc.; G is preferably D-glucose, D-galactose, etc.

[0042] The present invention provides a synthetic route for the anticancer drug TNBG-sugar coupling shown in Formula IV, as follows:

[0043]

[0044] The synthesis of the anticancer drug TNBG-sugar conjugate shown in Formula IV has the following characteristics:

[0045] The synthesis is carried out through cyclization reaction, chlorination reaction, and cyclization reaction to obtain the key intermediate 14, and then through substitution reaction, glycosylation reaction, and deacetylation reaction to obtain the target compound of formula IV;

[0046] The cyclization reaction is that the starting material reacts with diethyl oxalate under acidic conditions (dilute hydrochloric acid); the chlorination reaction is that phosphorus oxychloride is used to convert the hydroxyl group into a chlorine atom; the cyclization reaction is that under alkaline conditions (potassium carbonate) the cyclization reaction is carried out with 2-aminomethylpiperidine to form a four-membered ring key intermediate 14; the substitution reaction is that the hydrogen on the secondary amine is replaced by 1-bromoethanol or propanol under strong alkaline conditions (NaH (60%)); the glycosidation reaction is that the hydroxyl group reacts with a chirally fixed tetraacetyl brominated sugar under the catalysis of a heavy metal salt; the deacetylation reaction is carried out in a methanol solution in an alkaline environment (potassium carbonate) to obtain the target compounds IV-1 to IV-24.

[0047] In a third aspect, the present invention provides the following representative compounds or pharmaceutically acceptable salts thereof or stereoisomers thereof, etc.:

[0048]

[0049]

[0050] In a fourth aspect, the present invention provides specific uses of the compounds of Formula I and Formula IV or their pharmaceutically acceptable salts or stereoisomers thereof in the preparation of anti-tumor drugs;

[0051] The prepared anti-tumor drug is a drug for treating cancer, characterized in that: preferably, the cancer is lung cancer, liver cancer, etc.;

[0052] A medicine is a preparation prepared by using the compounds of formula I and formula IV or their pharmaceutically acceptable salts or stereoisomers as active ingredients and adding suitable excipients or auxiliary ingredients. BRIEF DESCRIPTION OF THE DRAWINGS

[0053] Figure 1 It is a schematic diagram of the general structure of the patent of this invention. DETAILED DESCRIPTION

[0054] Unless otherwise specified, the reagents and instruments used in the present invention are all commercially available, wherein the chemical reagents are all AR grade and do not require further purification. The present invention is now further described in detail in conjunction with the examples, but is not limited to these examples.

[0055] The structures of the compounds in the following examples are mainly determined by nuclear magnetic resonance (NMR) or high-resolution mass spectrometry (HRMS). NMR determination uses a Bruker AV-600 nuclear magnetic spectrometer (USA), with deuterated chloroform (CDCl3) or deuterated dimethyl sulfoxide (DMSO-d6) as solvent and tetramethylsilane (TMS) as internal standard; high-resolution mass spectrometry determination uses a Waters G2-S QTOF-MS mass spectrometer (USA); melting point data are collected by a WRS-2C melting point instrument (Shanghai, China).

[0056] In the first aspect, the synthesis of compounds of formula I:

[0057] Example 1: Synthesis of key intermediate 6

[0058] The synthetic route of key intermediate 6 is as follows:

[0059]

[0060] Reagents and conditions a are: chloroacetyl chloride, sodium carbonate, dichloromethane, 0°C;

[0061] Reagents and conditions b are: potassium phthalimide, N,N-dimethylformamide, 110°C;

[0062] Reagents and conditions c are: phosphorus pentoxide, zinc chloride, acetonitrile, 120°C;

[0063] Reagents and conditions d are: sodium triacetoxyborohydride, methanol, dichloromethane;

[0064] Reagents and conditions e are: hydrazine hydrate, methanol, dichloromethane, room temperature, concentrated hydrochloric acid;

[0065] The specific preparation method is as follows:

[0066] (1) Preparation of 2-chloro-N-phenethylacetamide (Compound 2)

[0067] Take a clean three-necked flask, add 60mL of dichloromethane and anhydrous sodium carbonate (14.4g, 0.135mol, 1.5eq), add chloroacetyl chloride (12.3mL, 0.154mol, 1.7eq) under ice-salt bath and stirring, slowly drop β-phenylethylamine (11g, 0.090mol, 1eq) dichloromethane solution, pay attention to control the temperature below 0℃, and monitor by TLC until the reaction is complete. Slowly pour the reaction solution into water, separate the water layer, extract with dichloromethane (2x 60mL) and saturated brine 20mL in turn, combine the organic layers, dry with anhydrous sodium sulfate, concentrate, and dry to obtain a light yellow solid (Compound 2), 16.87g, yield 94%.

[0068] (2) Preparation of 2-(1,3-dioxoisoindole-2-yl)-N-phenylacetamide (Compound 3)

[0069] Compound 2 (16.87 g, 0.085 mol, 1 eq) was dissolved in 70 mL of N,N-dimethylformamide, potassium phthalimide (20.55 g, 0.111 mol, 1.3 eq) was added under stirring, the temperature was raised to 110°C, the reaction was allowed to react for 5 h, a large amount of white solid was precipitated, filtered and dried to obtain a white solid (compound 3), 25.08 g, with a yield of 95.3%.

[0070] (3) Preparation of 2-((3,4-dihydroisoquinolin-1-yl)methyl)isoindole-1,3-dione (Compound 4)

[0071] Take a clean eggplant-shaped bottle, add compound 3 (25.08 g, 0.081 mol, 1 eq), phosphorus pentoxide (8.08 g, 0.057 mol, 0.7 eq), zinc chloride (3.88 g, 0.028 mol, 0.35 eq), phosphorus oxychloride (24.94 g, 0.163 mol, 2 eq) and 350 mL of acetonitrile in sequence, heat to 90 ° C with stirring, monitor by TLC until the reaction is complete, and concentrate under reduced pressure. Pour the reaction solution into ice water, adjust the pH value to weak alkalinity with alkaline water, precipitate a yellow solid, filter and dry to obtain a yellow solid (compound 4), 23.08 g, yield 97.8%.

[0072] (4) Preparation of 2-((1,2,3,4-tetrahydroisoquinolin-1-yl)methyl)isoindole-1,3-dione (Compound 5)

[0073] Take a clean eggplant-shaped bottle, add compound 4 (23.08g, 0.079mol, 1eq), 40mL methanol and 100mL dichloromethane, stir at room temperature, add sodium triacetoxyborohydride (67.4g, 0.318mol, 4eq) in four times, and monitor by TLC until the reaction is complete. Pour the reaction solution into water, adjust the pH value to alkaline with sodium carbonate, and some solids appear. Filter, wash the filter cake with dichloromethane, extract the water layer with dichloromethane (3x 100mL), dry with anhydrous sodium sulfate, concentrate, and dry to obtain a black solid (compound 5), 12.84g, yield 55.2%.

[0074] (5) Preparation of (1,2,3,4-tetrahydroisoquinolin-1-yl)methylamine hydrochloride (Compound 6)

[0075] Compound 5 (12.84 g, 0.044 mol, 1 eq) was dissolved in 100 mL of methanol, and 80% hydrated hydrazine (6.4 mL, 0.132 mol, 3 eq) was added at 80°C with stirring, and the reaction was monitored by TLC until it was complete. After standing and cooling, the flocculent was filtered, the filter cake was washed with methanol, saturated sodium bicarbonate solution was added to adjust to weak alkalinity, extracted with dichloromethane (3 x 100 mL), washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated to obtain a black oil (Compound 6), 5.51 g, yield 77.3%, which was directly used in the next step with Compound 9a.

[0076] Example 2: Synthesis of key intermediate 9

[0077] The synthetic route of key intermediate 9 is as follows:

[0078]

[0079] Reagents and conditions f are: diethyl oxalate, 4 mol / L hydrochloric acid, 90°C;

[0080] Reagents and conditions g were: phosphorus oxychloride, N,N-dimethylformamide, 90°C;

[0081] The specific preparation method is as follows:

[0082] (1) Preparation of 2,3-dihydroxyquinoxaline (Compound 8a)

[0083] Take a clean eggplant-shaped bottle, add o-phenylenediamine (10g, 0.092mol, 1eq), diethyl oxalate (20.27g, 0.139mol, 1.5eq) and 100mL of 4mol / L hydrochloric acid, stir, reflux at 90℃, react for 2h, monitor by TLC until the reaction is complete. Cool to room temperature, add 400mL of water, a large amount of solid precipitates, filter, wash with water, and dry to obtain a light green solid (compound 8a), 14.5g, yield 96.7%.

[0084] (2) Preparation of 2,3-dichloroquinoxaline (Compound 9a)

[0085] Take a dry eggplant-shaped bottle, add compound 8a (14.5g, 0.089mol, 1eq), 100mL of N,N-dimethylformamide and phosphorus oxychloride (25mL, 0.268mol, 3eq), stir, reflux at 90℃, react for 2h, monitor by TLC until the reaction is complete. Cool to room temperature, pour the reaction solution into crushed ice, stir until a large amount of solid precipitates, filter, wash with water, and dry to obtain a white solid (compound 9a), 16.17g, yield 90.9%.

[0086] Example 3: Synthesis of key intermediate 11

[0087] The synthetic route of key intermediate 11 is as follows:

[0088]

[0089] Reagents and conditions h are: potassium carbonate, N,N-dimethylformamide, 120°C;

[0090] Reagents and conditions i are: 3-bromopropanol, sodium hydride, N,N-dimethylformamide, 45°C;

[0091] The specific preparation method is as follows:

[0092] (1) Preparation of 4b,5,14,15-tetrahydro-6H-isoquinolino[2',1':1,6]pyrazino[2,3-b]quinoxaline (Compound 10)

[0093] Compound 6 (5.51 g, 0.034 mol, 1 eq) and compound 9a (6.76 g, 0.034 mol, 1 eq) were dissolved in 120 mL of N, N-dimethylformamide, potassium carbonate (9.79 g, 0.136 mol, 4 eq) was added, the temperature was raised to 110°C, refluxed for 12 h, and the reaction was monitored by TLC until it was complete. Then, the mixture was cooled to room temperature, a large amount of water was added, stirred, a large amount of solid was precipitated, filtered, and dried; recrystallized with a mixed solvent (isopropanol: tetrahydrofuran = 1:2), filtered, and dried to finally obtain a pink solid (compound 10), 3.31 g, with a yield of 33.8%.

[0094] (2) Preparation of 3-(4b,5,14,15-tetrahydro-6H-isoquinolin[2',1':1,6]pyrazino[2,3-b]quinoxaline-6-yl)-1-propanol (Compound 11)

[0095] Take a clean eggplant-shaped bottle, add compound 10 (1.96g, 0.007mol, 1eq), 120mL N,N-dimethylformamide, add sodium hydride (1.13g, 0.027mol, 4eq) under ice bath, stir for half an hour, add 3-bromopropanol (1.13g, 0.008mol, 1.2eq), reflux at 45°C, monitor by TLC until the reaction is complete. Cool to room temperature, add 120mL water, remove insoluble matter by suction, extract with dichloromethane (3x 100mL), wash with saturated brine 100mL, combine organic layers, dry over anhydrous sodium sulfate, concentrate, pass through column (mobile phase is dichloromethane), and concentrate to obtain a golden yellow solid (compound 11), 0.91g, yield 38.7%. 1 HNMR(600MHz,DMSO-d6)δ7.48-7.46(m,1H),7.46-7.43(m,1H),7.41-7.37(m,1H),7.30-7.23(m, 3H),7.22-7.17(m,2H),5.01-4.96(m,1H),4.86(dd,J=10.1,3.6Hz,1H),4.65(s,1H),4.10(dd,J= 11.9,3.7Hz,1H),3.88-3.82(m,1H),3.74-3.68(m,1H),3.50(t,J=6.2Hz,2H),3.49-3.45(m,1H) ,3.08(td,J=12.2,3.5Hz,1H),3.02-2.96m,1H),2.89(dt,J=15.9,3.0Hz,1H),1.89-1.81(m,2H). 13C NMR (151MHz, DMSO-d6) δ143.55,143.29,137.33,136.60,135.44,133.88,129.47,127.44,126. 80,126.16,125.28,124.84,124.68,124.01,58.94,52.67,51.90,45.49,39.64,28.89,28.52.

[0096] Example 4: Synthesis of five-membered ring (AE ring) TNBG-glucose / galactose conjugate

[0097] The synthetic route of the five-membered ring anticancer drug TNBG-sugar conjugate is as follows:

[0098]

[0099] Reagents and conditions j were: 2,3,4,6-tetraacetoxy-α-D-pyranose / galactose bromide, dichloromethane, silver(I) trifluoromethanesulfonate, 4A molecular sieves, room temperature;

[0100] Reagents and conditions k are: potassium carbonate, methanol, room temperature.

[0101] The specific preparation method is as follows:

[0102] (1) Preparation of (2R,3R,4S,5R,6R)-2-(acetoxymethyl)-6-(3-(4b,5,14,15-tetrahydro-6H-isoquinolin[2',1':1,6]pyrazin[2,3-b]quinoxalin-6-yl)propoxy)tetrahydro-2H-pyran-3,4,5-triacetate (Compound 12a)

[0103] Compound 11 (0.66 g, 1.91 mmol, 1 eq) was dissolved in 22 mL of dry dichloromethane, 3 g of 4A molecular sieves was added and stirred for 10 min, and 2,3,4,6-tetraacetoxy-α-D-pyranose glucopyranose bromide (1.25 g, 3.05 mmol, 1 eq) was added, nitrogen was protected, and stirred for 1 h. Silver (I) trifluoromethanesulfonate (0.78 g, 3.05 mmol, 1.6 eq) was added in an ice bath, nitrogen was protected, and the mixture was stirred at room temperature overnight. The reaction was monitored by TLC until the end. The mixture was filtered off with celite, concentrated, diluted with 20 mL of dichloromethane, washed successively with water (2 x 20 mL), saturated sodium bicarbonate solution (2 x 20 mL), and saturated brine 20 mL, dried over anhydrous sodium sulfate, concentrated, and passed through a column (mobile phase: petroleum ether: ethyl acetate = 1:1). The mixture was concentrated to obtain a yellow oil (compound 12a), 0.41 g, with a yield of 31.8%, and directly carried out to the next step.

[0104] (2) Preparation of 2R,3S,4S,5R,6R)-2-(acetoxymethyl)-6-(3-(4b,5,14,15-tetrahydro-6H-isoquinolin[2',1':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)tetrahydro-2H-pyran-3,4,5-triacetate (Compound 12b)

[0105] The preparation method was the same as that of Example 4-(1). The residue was concentrated to give a yellow oil (Compound 12b), 0.38 g, with a yield of 43.2%.

[0106] (3) Preparation of (2R,3R,4S,5R,6R)-2-(hydroxymethyl)-6-(3-(4b,5,14,15-tetrahydro-6H-isoquinolin[2',1':1,6]pyrazin[2,3-b]quinoxalin-6-yl)propoxy)tetrahydro-2H-pyran-3,4,5-triol (I-1)

[0107]

[0108] Add 15 mL of anhydrous methanol to compound 12a, adjust the pH to 9 with potassium carbonate, stir at room temperature, and monitor by TLC until the reaction is complete. Filter through diatomaceous earth, concentrate, and pass through a column (mobile phase is dichloromethane:methanol=30:1-10:1), and concentrate to obtain a light yellow solid (I-1), 80 mg, yield 26.0%, melting point 129.1-133.1°C. 1 H NMR (600MHz, DMSO-d6) δ7.53-7.46(m,1H),7.46-7.41(m,2H),7.30-7.23(m,3H),7.22-7.16(m,2H),5.06(dd,J=37.4,4 .8Hz,1H),5.01-4.97(m,1H),4.95(dd,J=4.9,1.6Hz,1H),4.91-4.87(m,2H),4.47(q,J=6.1Hz,1H),4.19(dd,J=10.9,7 .8Hz,1H),4.15-4.11(m,1H),3.94-3.89(m,1H),3.84-3.76(m,2H),3.68-3.63(m,1H),3.60-3.54(m,1H),3.52-3.47(m ,1H),3.45-3.37(m,1H),3.19-3.14(m,1H),3.12-2.96(m,5H),2.92-2.87(m,1H),2.07-1.99(m,1H),1.99-1.91(m,1H). 13CNMR(151MHz,DMSO-d6)δ143.57,143.17,137.47,136.66,135.44,133.96,129.43,127.40,126.83,126.21,125.25,125.06,124 .61,124.01,103.41,77.42,77.20,74.05,70.69,67.02,66.60,61.63,52.72,52.17,45.87,28.54,26.94.HRMS(ESI):m / z[M+H] + Calcd for C 27 H 33 N4O6:509.2400; Found:509.2402.

[0109] (4) Preparation of (2R,3S,4S,5R,6R)-2-(hydroxymethyl)-6-(3-(4b,5,14,15-tetrahydro-6H-isoquinolin[2',1':1,6]pyrazin[2,3-b]quinoxalin-6-yl)propoxy)tetrahydro-2H-pyran-3,4,5-triol (I-2)

[0110]

[0111] The preparation method was the same as that of Example 4-(3). A pale yellow solid (I-2) was obtained by column chromatography, 110 mg, yield 38.5%, melting point 101.0-103.9°C. 1 HNMR(600MHz,DMSO-d6)δ7.53-7.50(m,1H),7.46-7.40(m,2H),7.30-7.23(m,3H),7.22-7.16(m,2H),5.02-4.97(m,1H),4 .94-4.85(m,2H),4.71(t,J=5.3Hz,1H),4.55-4.49(m,1H),4.36(dd,J=30.6,4.5Hz,1H),4.16-4.10(m,2H),3.93-3.86(m, 1H),3.84-3.72(m,2H),3.65(q,J=3.5Hz,1H),3.59-3.51(m,2H),3.51-3.44(m,2H),3.3-3.33(m,2H),3.31-3.27(m,1H), 3.12-3.05(m,1H),2.99(td,J=13.7,11.5,5.3Hz,1H),2.89(dt,J=15.9,3.0Hz,1H),2.07-1.99(m,1H),1.98-1.90(m,1H). 13C NMR(151MHz,DMSO-d6)δ143.56,143.17,137.47,136.65,135.33,133.93,129.40,127.40,126.93,126.49,126.30,125.24,124 .59,123.99,103.89,75.75,73.95,71.11,68.61,66.84,66.45,60.87,52.69,52.30,45.65,28.53,26.92.HRMS(ESI):m / z[M+H] + Calcd for C 27 H 33 N4O6:509.2400; Found:509.2402.

[0112] In the second aspect, the synthesis of the compound of formula IV:

[0113] Example 5: Synthesis of key intermediate 15

[0114] The synthetic route of key intermediate 15 is as follows:

[0115]

[0116] Reagents and conditions h are: potassium carbonate, N,N-dimethylformamide, 120°C;

[0117] Reagents and conditions i are: 3-bromopropanol, sodium hydride, N,N-dimethylformamide, 45°C;

[0118] The specific preparation method is as follows:

[0119] (1) Preparation of 2,3,4,4a,5,6-hexahydro-1H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxaline (Compound 14a)

[0120] Dissolve compound 9a (16.60 g, 0.083 mol, 1 eq) in 200 mL N, N-dimethylformamide, add compound 13 (14.29 g, 0.125 mol, 1.5 eq) and potassium carbonate (34.58 g, 0.250 mol, 3 eq) in sequence, heat to 110°C, reflux for 3 h, monitor by TLC until the reaction is complete. Cool to room temperature, add a large amount of water, stir, precipitate a large amount of solid, filter, and dry; recrystallize with a mixed solvent (dichloromethane: methanol = 1:2), filter, and dry to finally obtain a light yellow solid (compound 14a), 19.99 g, yield 99.8%. 1 H NMR (600 MHz, Chloroform- d)δ7.58-7.54(m,1H),7.44-7.39(m,1H),7.28-7.22(m,2H),6.65(s,1H),4.95-4.89(m,1H),3.57-3.50(m,1H),3.41-3.32(m,2H),2.7 2(td,J=13.0,3.1Hz,1H),1.91-1.84(m,2H),1.81-1.77(m,1H),1.62-1.54(m,4.3,3.0Hz,1H),1.53-1.45(m,1H),1.41-1.34(m,1H). 13 C NMR (151MHz, Chloroform-d) δ144.24,144.17,137.77,136.78,125.68,124.69,124.30,124.06,52.99,45.35,44.33,30.09,24.53,23.38.

[0121] (2) Preparation of 9-chloro-2,3,4,4a,5,6-hexahydro-1H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxaline (Compound 14f)

[0122] The preparation method was the same as that of Example 5-(1). A pale purple solid (Compound 14f) was obtained by column chromatography with a yield of 40.8%. 1 H NMR(600MHz,Chloroform-d)δ7.46(d,J=8.7Hz,1H),7.40(d,J=2.4Hz,1H),7.1 9(dd,J=8.7,2.4Hz,1H),5.96(s,1H),4.91-4.86(m,1H),3.57-3.52(m,1H),3.4 2-3.36(m,2H),2.73(td,J=13.0,3.0Hz,1H),1.94-1.90(m,1H),1.89-1.85(m,1 H),1.84-1.80(m,1H),1.62-1.55(m,1H),1.55-1.47(m,1H),1.43-1.36(m,1H). 13 C NMR (151MHz, Chloroform-d) δ144.16,144.08,137.25,136.28,129.68,126.58,124.84,123.44,52.91,45.34,44.31,30.05,24.46,23.30.

[0123] (3) The preparation method of intermediates 14b-14e is the same as that of "Example 5-(1)".

[0124] (4) Preparation of 2-(1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxaline-6-yl)-1-ethanol (Compound 15a)

[0125] Compound 14a (2.50 g, 0.010 mol, 1 eq) and 70 mL of N,N-dimethylformamide were added to a clean eggplant-shaped bottle, and 60% sodium hydride (1.66 g, 0.042 mol, 4 eq) was added under ice bath. After stirring for half an hour, 2-bromoethanol (1.95 g, 0.016 mol, 1.5 eq) was added, and refluxed at 45°C. TLC was monitored until the reaction was completed. After cooling to room temperature, 100 mL of water was added, the insoluble matter was removed by suction, and the mixture was washed with dichloromethane (3 x 100 mL) and saturated brine. The organic layers were combined, dried over anhydrous sodium sulfate, concentrated, and passed through a column (the mobile phase was dichloromethane). The mixture was concentrated to obtain a light yellow solid (compound 15a), 1.80 g, with a yield of 60.9%. 1 H NMR(600MHz,Chloroform-d)δ7.56-7.52(m,1H),7.50-7.45m,1H),7.25-7.22(m ,2H),5.42(s,1H),4.87-4.83m,1H),3.96-3.89m,2H),3.83-3.76(m,2H),3.46( d,J=6.3Hz,2H),3.39-3.33(m,1H),2.70(td,J=13.0,3.1Hz,1H),1.93-1.84(m, 2H),1.82-1.77(m,1H),1.62-1.53(m,1H),1.53-1.45(m,1H),1.38-1.30m,1H). 13 C NMR(151MHz,Chloroform-d)δ144.29,144.29,137.15,135.99,125.43,124. 87,124.51,124.34,62.67,53.67,53.36,52.38,44.44,29.93,24.39,23.22.

[0126] (5) Preparation of 3-(1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxaline-6-yl)-1-propanol (Compound 15b)

[0127] The preparation method was the same as that of Example 5-(4). A pale yellow solid (Compound 15b) was obtained by column chromatography with a yield of 74.1%. 1H NMR(600MHz,Chloroform-d)δ7.56-7.52(m,1H),7.48-7.44(m,1H),7.26-7.21m,2H) ,5.48(s,1H),4.88-4.82(m,1H),3.90-3.85(m,1H),3.80-3.74(m,1H),3.58-3.53(m ,1H),3.53-3.48(m,1H),3.43-3.38(m,1H),3.37-3.30(m,2H),2.71(td,J=13.1,3.1 Hz,1H),1.94-1.78(m,5H),1.64-1.56(m,1H),1.54-1.46(m,1H),1.39-1.32(m,1H). 13 CNMR(151MHz,Chloroform-d)δ143.88,143.73,136.80,136.26,125.50,125.06, 124.26,124.07,57.66,52.07,51.33,44.66,44.38,30.18,29.99,24.31,23.25.

[0128] (6) The preparation method of intermediates 15c-151 is the same as that of "Example 5-(4)".

[0129] Example 6: Synthesis of Four-membered Ring (ABCE Ring) TNBG-Glucose / Galactose Conjugate

[0130] The synthetic route of the four-membered ring (ABCE ring) anticancer drug TNBG-sugar conjugate is as follows:

[0131]

[0132] Reagents and conditions j were: 2,3,4,6-tetraacetoxy-α-D-pyranose / galactose bromide, dichloromethane, silver(I) trifluoromethanesulfonate, 4A molecular sieves, room temperature;

[0133] Reagents and conditions k are: potassium carbonate, methanol, room temperature.

[0134] The specific preparation method is as follows:

[0135] (1) Preparation of (2R,3R,4S,5R,6R)-2-(acetoxymethyl)-6-(2-(1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)tetrahydro-2H-pyran-3,4,5-triacetate (Compound 16a)

[0136] Compound 15a (1.3 g, 4.57 mmol, 1 eq) was dissolved in 18 mL of dry dichloromethane, 0.36 g of activated 4A molecular sieve was added and stirred for 10 min, and 2,3,4,6-tetraacetoxy-α-D-pyranose glucopyranose bromide (2.26 g, 5.49 mmol, 1.2 eq) was added, nitrogen was protected, and after stirring for 1 h, silver (I) trifluoromethanesulfonate (1.41 g, 5.49 mmol, 1.2 eq) was added in an ice bath, nitrogen was protected, and the mixture was stirred at room temperature overnight. The reaction was monitored by TLC until the end. The mixture was filtered through celite, concentrated, diluted with 20 mL of dichloromethane, washed successively with water (2 x 20 mL), saturated sodium bicarbonate solution (2 x 20 mL), and saturated brine 20 mL, dried over anhydrous sodium sulfate, concentrated, and passed through a column (mobile phase: petroleum ether: ethyl acetate = 1:1) to obtain a light yellow solid (compound 16a), 0.63 g, yield 22.4%, which was directly used for the next step.

[0137] (2) Preparation of (2R,3S,4S,5R,6R)-2-(acetoxymethyl)-6-(3-(9-chloro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)tetrahydro-2H-pyran-3,4,5-triacetate (Compound 16x)

[0138] The preparation method was the same as that of Example 6-(1). A pale yellow solid (Compound 16x) was obtained by column chromatography. The product was 0.90 g with a yield of 86.5%. The product was directly used for the next step.

[0139] (3) The preparation method of compounds 16b-16w is the same as that of "Example 6-(1)".

[0140] (4) Preparation of (2R,3R,4S,5R,6R)-2-(2-(1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-1)

[0141]

[0142] Compound 16a (0.63 g, 1.02 mmol, 1 eq) was dissolved in 15 mL of anhydrous methanol, and the pH was adjusted to 9 with potassium carbonate, and the reaction was stirred at room temperature and monitored by TLC until the reaction was completed. The mixture was filtered through diatomaceous earth, concentrated, and passed through a column (the mobile phase was dichloromethane: methanol = 30:1-10:1). The mixture was concentrated to obtain a white solid (IV-1), 150 mg, with a yield of 32.8%, and a melting point of 94.1-96.0°C. 1H NMR (600MHz, DMSO-d6) δ7.39(dd,J=12.8,7.8Hz,2H),7.21-7.13(m,2H),5.03(t,J=5.1Hz,1H),4.93(t,J=4.4Hz,1H ),4.88(d,J=5.3Hz,1H),4.68(d,J=13.3Hz,1H),4.48-4.41(m,1H),4.19(d,J=7.8Hz,1H),4.04-3.97(m,1H),3.94- 3.86(m,1H),3.79-3.71(m,2H),3.69-3.57(m,2H),3.54-3.39(m,3H),3.15-3.12(m,1H),3.11-3.07(m,1H),3.07-3 .02(m,1H),2.97(q,J=7.5Hz,1H),2.67(t,J=12.1Hz,1H),1.84-1.76(m,3H),1.50-1.42(m,2H),1.30-1.25(m,1H). 13 C NMR(151MHz,DMSO-d6)δ144.32,143.23,137.32,136.82,125.37,124.93,124.63,123.94,103.64,77.36, 77.17,73.95,70.55,66.65,61.55,52.15,51.89,47.89,44.44,29.61,24.44,23.22.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 31 N4O6:447.2244; Found:447.2245.

[0143] (5) Preparation of (2R,3S,4S,5R,6R)-2-(2-(1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-2)

[0144]

[0145] The preparation method was the same as that of Example 6-(4). A white solid (IV-2) was obtained by column chromatography, 60 mg, with a yield of 18.4% and a melting point of 51.8-54.1°C. 1H NMR(600MHz,DMSO-d6)δ7.42-7.36(m,2H),7.20-7.14(m,2H),4.89-4.86m,1H),4.70-4.65(m,2H),4. 54-4.50(m,1H),4.35-4.32(m,1H),4.13(d,J=7.4Hz,1H),4.02-3.96(m,1H),3.92-3.84(m,1H),3.81

[0146] -3.64(m,3H),3.64-3.61(m,1H),3.54-3.41(m,3H),3.39-3.34(m,2H),3.31-3.28(m,1H),3. 28-3.24(m,1H),2.71-2.63(m,1H),1.84-1.76(m,3H),1.50-1.41(m,2H),1.29-1.22(m,1H). 13 C NMR(151MHz,DMSO-d6)δ144.32,143.20,137.32,136.81,125.37,124.93,124.62,123.91,104.25,75.65, 73.89,71.06,68.59,66.52,60.83,52.11,51.91,47.95,44.42,29.62,24.43,23.21.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 31 N4O6:447.2244; Found:447.2248.

[0147] (6) Preparation of (2R,3R,4S,5R,6R)-2-(3-(1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-(6-hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-3)

[0148]

[0149] The preparation method was the same as that of Example 6-(4). A white solid (IV-3) was obtained by column chromatography, 450 mg, with a yield of 43.9% and a melting point of 105.6-108.3°C. 1H NMR(600MHz,DMSO-d6)δ7.39(d,J=7.6Hz,2H),7.21-7.12(m,2H),5.02(dd,J=12.9,4.8Hz,1H),4.99-4.86(m,2H),4.68 (d,J=13.3Hz,1H),4.48(t,J=6.0Hz,1H),4.15(dd,J=7.8,2.9Hz,1H),3.91-3.81(m,1H),3.72-3.64(m,3H),3.63-3.57( m,1H),3.56-3.50(m,1H),3.46-3.41(m,1H),3.40-3.36(m,2H),3.15(t,J=8.8Hz,1H),3.12-3.08(m,1H),3.08-3.03(m ,1H),3.02-2.95(m,1H),2.71-2.63(m,1H),1.98-1.87(m,2H),1.84-1.75(m,3H),1.50-1.41(m,2H),1.29-1.24(m,1H). 13 C NMR (151MHz, DMSO-d6) δ144.27,143.34,137.45,136.77,125.33,124.94,124.57,123.85,103.32,77.35,77. 19,73.97,70.58,66.77,61.61,52.15,51.12,45.25,44.40,29.68,26.87,24.40,23.23.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 33 N4O6:461.2400; Found:461.2403.

[0150] (7) Preparation of (2R,3S,4S,5R,6R)-2-(3-(1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-4)

[0151]

[0152] The preparation method was the same as that of Example 6-(4). A pale yellow solid (IV-4) was obtained by column chromatography, 115 mg, with a yield of 39.2% and a melting point of 150.2-152.9°C. 1HNMR(600MHz,DMSO-d6)δ7.42-7.36(m,2H),7.21-7.11(m,2H),4.86(dd,J=11.9,4.6Hz,1H),4.70(d,J=5.4Hz,1 H),4.70-4.65(m,1H),4.58-4.52(m,1H),4.35(dd,J=4.6,2.8Hz,1H),4.10(dd,J=7.4,3.8Hz,1H),3.86-3.80(m, 1H),3.75-3.59(m,4H),3.56-3.49(m,2H),3.49-3.45(m,1H),3.41-3.34(m,3H),3.32-3.29(m,1H),3.29-3.26( m,1H),2.67(td,J=12.8,3.2Hz,1H),1.97-1.86(m,2H),1.85-1.75(m,3H),1.51-1.42(m,2H),1.30-1.24(m,1H). 13 C NMR (151MHz, DMSO-d6) δ144.26,143.34,137.46,136.76,125.33,124.95,124.57,123.83,103.81,75.65,73. 92,71.06,68.59,66.37,60.89,52.16,51.20,45.25,44.39,29.66,26.93,24.41,23.23.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 33 N4O6:461.2400; Found:461.2404.

[0153] (8) Preparation of (2R,3R,4S,5R,6R)-2-(2-(9,10-dimethyl-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-5)

[0154]

[0155] The preparation method was the same as that of Example 6-(4). A yellow solid (IV-5) was obtained by column chromatography, 160 mg, with a yield of 27.8% and a melting point of 159.6-162.4°C. 1HNMR (600MHz, DMSO-d6) δ7.20 (s, 1H), 7.19 (s, 1H), 5.03 (d, J = 5.8Hz, 1H), 4.99-4.91 (m, 1H), 4.91-4.86 (m, 1H), 4. 65-4.60(m,1H),4.46(d,J=11.9Hz,1H),4.18(d,J=7.8Hz,1H),4.03-3.96(m,1H),3.90-3.80(m,1H),3.80-3.69(m, 2H),3.69-3.57(m,2H),3.49-3.40(m,2H),3.32-3.28(m,1H),3.14(t,J=8.8Hz,1H),3.11-3.07(m,1H),3.07-3.02( m,1H),2.99-2.94(m,1H),2.66-2.59(m,1H),2.26(s,6H),1.83-1.74(m,3H),1.48-1.39(m,2H),1.27-1.22(m,1H). 13 C NMR(151MHz,DMSO-d6)δ143.96,142.85,135.46,134.96,133.42,132.63,125.36,124.92,103.47,77.10,76.84, 73.77,70.36,66.65,61.37,55.20,52.17,47.81,44.45,29.57,24.38,23.16,19.79,19.76.HRMS(ESI):m / z[M+H] + Calcd for C 24 H 35 N4O6:475.2557; Found:475.2557.

[0156] (9) Preparation of (2R,3S,4S,5R,6R)-2-(2-(9,10-dimethyl-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-6)

[0157]

[0158] The preparation method was the same as that of Example 6-(4). A pale yellow solid (IV-6) was obtained by column chromatography, 140 mg, with a yield of 23.4% and a melting point of 216.6-219.1°C. 1HNMR(600MHz,DMSO-d6)δ7.20(s,1H),7.19(s,1H),4.87(t,J=4.4Hz,1H),4.71-4.66(m,1H),4.66-4 .60(m,1H),4.55-4.49(m,1H),4.36-4.32(m,1H),4.12(dd,J=7.4,1.1Hz,1H),4.01-3.94(m,1H),3. 88-3.80(m,1H),3.79-3.72(m,1H),3.72-3.67(m,1H),3.64-3.57(m,2H),3.54-3.40(m,3H),3.33-3 .24(m,4H),2.66-2.59(m,1H),2.26(s,6H),1.82-1.74(m,3H),1.48-1.39(m,2H),1.27-1.19(m,1H). 13 C NMR(151MHz,DMSO-d6)δ143.95,142.85,135.59,135.05,133.29,132.47,125.47,125.05,104.25,75.65,73.87, 71.07,68.59,66.51,60.83,55.38,52.15,47.94,44.38,29.58,24.47,23.23,19.83,19.80.HRMS(ESI):m / z[M+H] + Calcd for C 24 H 35 N4O6:475.2557; Found:475.2560.

[0159] (10) Preparation of (2R,3R,4S,5R,6R)-2-(3-(9,10-dimethyl-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-7)

[0160]

[0161] The preparation method was the same as that of Example 6-(4). A white solid (IV-7) was obtained by column chromatography, 190 mg, with a yield of 42.6% and a melting point of 116.8-119.2°C. 1HNMR(600MHz,DMSO-d6)δ7.21(s,1H),7.19(s,1H),5.00(dd,J=13.6,4.8Hz,1H),4.97-4.83(m,2H),4.64(d,J=1 2.7Hz,1H),4.50-4.41(m,1H),4.15(dd,J=7.8,2.6Hz,1H),3.88-3.81(m,1H),3.70-3.63(m,3H),3.63-3.48(m, 3H),3.47-3.48(m,2H),3.17-3.13(m,1H),3.12-3.07(m,1H),3.07-3.02(m,1H),2.97(t,J=8.7Hz,1H),2.66-2. 60(m,1H),2.26(s,3H),2.25(s,3H),1.95-1.84(m,2H),1.82-1.75(m,3H),1.48-1.42(m,2H),1.29-1.23(m,1H). 13 C NMR (151MHz, DMSO-d6) δ143.87,143.01,135.75,135.03,133.22,132.38,125.45,125.08,103.21,77.33,77.21,73. 97,70.59,66.65,61.61,52.18,51.27,45.24,44.36,29.60,26.87,24.45,23.26,19.82,19.80.HRMS(ESI):m / z[M+H] + Calcd for C 25 H 37 N4O6:489.2713; Found:489.2717.

[0162] (11) Preparation of (2R,3S,4S,5R,6R)-2-(3-(9,10-dimethyl-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-8)

[0163]

[0164] The preparation method was the same as that of Example 6-(4). A white solid (IV-8) was obtained by column chromatography, 50 mg, yield 14.6%, melting point 186.9-190.0°C. 1HNMR(600MHz,DMSO-d6)δ7.20(s,1H),7.19(s,1H),4.83(s,1H),4.73-4.61(m,2H),4.54(s,1H) ),4.38-4.28(m,1H),4.10(dd,J=7.3,3.4Hz,1H),3.85-3.80(m,1H),3.70-3.60(m,3H),3.60-3 .56(m,1H),3.55-3.50(m,2H),3.49-3.45(m,1H),3.32-3.25(m,5H),2.66-2.60(m,1H),2.26(s ,3H),2.25(s,3H),1.95-1.85(m,2H),1.83-1.75(m,3H),1.50-1.40(m,2H),1.29-1.24(m,1H). 13 C NMR (151MHz, DMSO-d6) δ143.89,142.99,135.55,134.88,133.36,132.55,125.31,124.93,103.78,75.46,73.68,70. 95,68.47,66.81,60.80,52.22,51.24,45.20,44.44,29.62,26.85,24.37,23.18,19.77,19.74.HRMS(ESI):m / z[M+H] + Calcd for C 25 H 37 N4O6:489.2713; Found:489.2716.

[0165] (12) Preparation of (2R,3R,4S,5R,6R)-2-(2-(9,10-difluoro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-9)

[0166]

[0167] The preparation method was the same as that of Example 6-(4). A pale yellow solid (IV-9) was obtained by column chromatography, 100 mg, yield 14.9%, melting point 114.8-117.6°C. 1HNMR (600MHz, DMSO-d6) δ7.37-7.33(m,1H),7.33-7.29(m,1H),5.03(t,J=4.7Hz,1H),4.94(t,J=4.3Hz,1H),4.89(d,J=5.3Hz ,1H),4.62(d,J=13.3Hz,1H),4.48-4.42(m,1H),4.18(d,J=7.8Hz,1H),4.02-3.95(m,1H),3.92-3.84(m,1H),3.81-3.75(m,1 H),3.74-3.70(m,1H),3.70-3.62(m,2H),3.52-3.43(m,1H),3.43-3.40(m,1H),3.39-3.36(m,1H),3.15-3.11(m,1H),3.11-3 .07(m,1H),3.06-3.01(m,1H),2.99-2.93(m,1H),2.71-2.64(m,1H),1.84-1.74m,3H),1.49-1.40(m,2H),1.28-1.21(m,1H). 13 CNMR(151MHz,DMSO-d6)δ147.95,146.36,144.49,143.39,133.94,133.37,111.61,111.15,103.46,77.36, 77.15,73.93,70.55,66.51,61.57,52.00,51.70,47.95,44.34,29.55,24.36,23.11.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 29 N4O6F2:483.2055; Found:483.2059.

[0168] (13) Preparation of (2R,3S,4S,5R,6R)-2-(2-(9,10-difluoro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-10)

[0169]

[0170] The preparation method was the same as that of Example 6-(4). A white solid (IV-10) was obtained by column chromatography, 100 mg, yield 16.4%, melting point 245.3-248.0°C. 1HNMR(600MHz,DMSO-d6)δ7.36-7.33(m,1H),7.33-7.28(m,1H),4.87(t,J=4.4Hz,1H),4.74-4.66(m,1H),4.65-4.60(m,1H) ,4.52(t,J=5.7Hz,1H),4.37-4.30(m,1H),4.13(d,J=7.3Hz,1H),4.01-3.94(m,1H),3.90-3.82(m,1H),3.79-3.74(m,1H),3 .74-3.70(m,1H),3.70-3.65(m,1H),3.64-3.61(m,1H),3.54-3.46(m,2H),3.45-3.41(m,1H),3.40-3.35(m,1H),3.32-3.3 0(m,1H),3.30-3.27(m,1H),3.27-3.24(m,1H),2.71-2.64(m,1H),1.83-1.75(m,3H),1.49-1.40(m,2H),1.29-1.20(m,1H). 13 C NMR(151MHz,DMSO-d6)δ147.90,146.30,144.48,143.38,133.74,133.21,111.56,111.10,103.79,75.39, 73.54,70.87,68.43,66.29,60.73,51.91,51.65,47.96,44.41,29.50,24.26,22.94.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 29 N4O6F2:483.2055; Found:483.2059.

[0171] (14) Preparation of (2R,3R,4S,5R,6R)-2-(3-(9,10-difluoro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-11)

[0172]

[0173] The preparation method was the same as that of Example 6-(4). A white solid (IV-11) was obtained by column chromatography, 90 mg, with a yield of 17.3% and a melting point of 163.4-165.8°C. 1HNMR (600MHz, DMSO-d6) δ7.36-7.28(m,2H),5.01(dd,J=12.9,4.9Hz,1H),4.94(dd,J=4.8,1.0Hz,1H),4.89(d,J=5.1Hz,1H),4. 63(dd,J=13.2,2.6Hz,1H),4.46(t,J=5.8Hz,1H),4.14(dd,J=7.8,3.7Hz,1H),3.87-3.82(m,1H),3.73-3.66(m,2H),3.66-3.63 (m,1H),3.63-3.59(m,1H),3.54-3.49(m,1H),3.46-3.38(m,2H),3.38-3.34(m,1H),3.17-3.12(m,1H),3.11-3.07(m,1H),3.06 -3.02(m,1H),3.00-2.95(m,1H),2.71-2.65(m,1H),1.96-1.84(m,2H),1.83-1.75(m,3H),1.50-1.40(m,2H),1.29-1.22(m,1H). 13 CNMR(151MHz,DMSO-d6)δ147.89,146.30,144.46,143.54,134.08,133.32,111.67,111.26,103.30,77.36,77. 20,73.97,70.60,66.66,61.62,52.00,50.93,45.30,44.37,29.64,26.81,24.33,23.13.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 31 N4O6F2:497.2212; Found:497.2215.

[0174] (15) Preparation of (2R,3S,4S,5R,6R)-2-(3-(9,10-difluoro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-12)

[0175]

[0176] The preparation method was the same as that of Example 6-(4). A white solid (IV-12) was obtained by column chromatography, 92 mg, yield 20.5%, melting point 108.2-111.2°C. 1HNMR(600MHz,DMSO-d6)δ7.35-7.30(m,2H),4.86(dd,J=12.4,4.6Hz,1H),4.70(d,J=5.4Hz,1H) ,4.66-4.60(m,1H),4.56-4.52(m,1H),4.34(dd,J=4.6,2.9Hz,1H),4.09(dd,J=7.4,4.3Hz,1H) ,3.86-3.79(m,1H),3.74-3.59(m,4H),3.55-3.44(m,3H),3.42-3.35(m,2H),3.33-3.25(m,3H) ,2.71-2.65m,1H),1.95-1.85(m,2H),1.84-1.76(m,3H),1.51-1.40(m,2H),1.28-1.22(m,1H). 13 C NMR (151MHz, DMSO-d6) δ147.85,146.16,144.40,143.49,133.87,133.06,111.39,111.08,103.62,75.35,73. 56,70.93,68.45,66.43,60.79,51.95,50.86,45.28,44.45,29.54,26.67,24.22,22.98.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 30 N4O6F2Na:519.2031; Found:519.2033.

[0177] (16) Preparation of (2R,3R,4S,5R,6R)-2-(2-(9,10-dichloro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-13)

[0178]

[0179] The preparation method was the same as that of Example 6-(4). A yellow solid (IV-13) was obtained by column chromatography, 210 mg, with a yield of 34.4% and a melting point of 141.2-144.1°C. 1HNMR(600MHz,DMSO-d6)δ7.54(s,1H),7.52(s,1H),5.03(dd,J=5.1,3.3Hz,1H),4.94(dd,J=4.9,3.7Hz,1H) ,4.89(dd,J=5.3,1.0Hz,1H),4.67-4.62(m,1H),4.47-4.43(m,1H),4.18(dd,J=7.8,1.1Hz,1H),4.02-3.96( m,1H),3.93-3.86(m,1H),3.82-3.73(m,2H),3.73-3.63(m,2H),3.46-3.38(m,3H),3.15-3.07(m,2H),3.06- 3.01(n,1H),2.98-2.93(m,1H),2.74-2.66(m,1H),1.85-1.76(m,3H),1.48-1.41(m,2H),1.28-1.23(m,1H). 13 CNMR(151MHz,DMSO-d6)δ144.88,143.78,136.85,136.44,125.97,125.61,125.26,125.21,103.41,77.12, 76.82,73.74,70.37,66.49,61.38,51.99,51.57,48.08,44.43,29.49,24.29,22.97.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 29 N4O6Cl2:515.1464; Found:515.1468.

[0180] (17) Preparation of (2R,3S,4S,5R,6R)-2-(2-(9,10-dichloro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-14)

[0181]

[0182] The preparation method was the same as that of Example 6-(4). A pale yellow solid (IV-14) was obtained by column chromatography, 110 mg, with a yield of 22.4% and a melting point of 206.8-209.7°C. 1HNMR (600MHz, DMSO-d6) δ7.54(s,1H),7.52(s,1H),4.87(dd,J=4.8,3.3Hz,1H),4.69(dd,J=5.3,1.5Hz,1 H),4.67-4.62(m,1H),4.53-4.50(m,1H),4.34(t,J=4.3Hz,1H),4.13(d,J=7.3Hz,1H),4.02-3.95(m,1H) ,3.89-3.83(m,1H),3.80-3.67(m,3H),3.64-3.60(m,1H),3.54-3.45(m,2H),3.44-3.39(m,2H),3.32-3. 27(m,2H),3.27-3.24(m,1H),2.73-2.66(m,1H),1.84-1.76(m,3H),1.49-1.40(m,2H),1.29-1.23(m,1H). 13 CNMR(151MHz,DMSO-d6)δ144.91,143.80,136.97,136.52,125.95,125.70,125.29,125.21,104.18,75.63, 73.85,71.04,68.56,66.34,60.80,51.97,51.67,48.16,44.37,29.53,24.34,23.04.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 28 N4O6Cl2Na:537.1284; Found:537.1288.

[0183] (18) Preparation of (2R,3R,4S,5R,6R)-2-(3-(9,10-dichloro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-15)

[0184]

[0185] The preparation method was the same as that of Example 6-(4). A pale yellow solid (IV-15) was obtained by column chromatography, 260 mg, with a yield of 31.3% and a melting point of 145.7-148.5°C. 1H NMR (600MHz, DMSO-d6) δ7.54(d,J=1.3Hz,2H),5.03(dd,J=14.0,4.9Hz,1H),4.97-4.93(m,1H),4.91(d,J=5. 2Hz,1H),4.68-4.63(m,1H),4.47(t,J=5.9Hz,1H),4.14(dd,J=7.8,4.0Hz,1H),3.87-3.81(m,1H),3.74-360 (m,4H),3.54-3.49(m,1H),3.48-3.39(m,3H),3.17-3.12(m,1H),3.11-3.07(m,1H),3.06-3.01(m,1H),3.00 -2.95(m,1H),2.73-2.68(m,1H),1.95-1.86(m,2H),1.85-1.76(m,3H),1.49-1.41(m,2H),1.29-1.24(m,1H). 13 C NMR (151MHz, DMSO-d6) δ144.84,143.95,137.11,136.50,125.91,125.65,125.31,125.14,103.31,77.36,77. 19,73.96,70.59,66.65,61.61,52.00,50.85,45.48,44.37,29.64,26.78,24.33,23.06.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 31 N4O6Cl2:529.1621; Found:529.1622.

[0186] (19) Preparation of (2R,3S,4S,5R,6R)-2-(3-(9,10-dichloro-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-16)

[0187]

[0188] The preparation method was the same as that of Example 6-(4). A yellow solid (IV-16) was obtained by column chromatography, 40 mg, with a yield of 11.8% and a melting point of 106.8-109.6°C. 1H NMR(600MHz,DMSO-d6)δ7.53(d,J=0.9Hz,2H),4.85(dd,J=13.4,4.6Hz,1H),4.70(d,J=5.3 Hz,1H),4.66(dd,J=13.8,3.5Hz,1H),4.55-4.52(m,1H),4.34(dd,J=4.6,2.9Hz,1H),4.10( dd,J=7.4,4.4Hz,1H),3.85-3.80(m,1H),3.76-3.59(m,4H),3.56-3.49(m,2H),3.49-3.42 (m,2H),3.40-3.35(m,1H),3.34(m,1H),3.31-3.25(m,2H),2.74-2.68(m,1H),1.96-1.85(m 2H),1.85-1.76(m,3H),1.51-1.40(m,2H),1.30-1.23(m,1H). 13 C NMR (151MHz, DMSO-d6) δ144.87,143.94,137.12,136.49,125.91,125.66,125.30,125.12,103.94,75.66,73. 93,71.06,68.59,66.57,60.90,52.01,50.92,45.49,44.35,29.56,26.83,24.33,23.05.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 30 N4O6Cl2Na:551.1440; Found:551.1443.

[0189] (20) Preparation of (2R,3R,4S,5R,6R)-2-(hydroxymethyl)-6-(2-(9-methoxy-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)tetrahydro-2H-pyran-3,4,5-triol (IV-17)

[0190]

[0191] The preparation method was the same as that of Example 6-(4). A white solid (IV-17) was obtained by column chromatography, 88 mg, with a yield of 17.8% and a melting point of 181.9-183.9°C. 1HNMR (600MHz, DMSO-d6) δ7.32(dd,J=8.8,0.8Hz,1H),6.87(d,J=2.8Hz,1H),6.80(dd,J=8.8,2.8Hz,1H),5.04(t,J=4.7Hz,1H),4.94(t,J=4. 4Hz,1H),4.89(dd,J=5.3,1.5Hz,1H),4.60-4.55(m,1H),4.47-4.43(m,1H),4.19(dd,J=7.8,1.6Hz,1H),4.04-3.97(m,1H),3.95-3.84(m,1H ),3.82-3.79(m,1H),3.78(s,3H),3.77-3.73(m,1H),3.73-3.67(m,1H ),3.67-3.62(m,1H),3.52-3.40(m,2H),3.31-3.25(m,1H),3.16-3.11 (m,1H),3.11-3.07(m,1H),3.07-3.02(m,1H),2.99-2.95(m,1H),2.65 -2.57(m,1H),1.86-1.72(m,3H),1.50-1.38(m,2H),1.28-1.19(m,1H). 13 C NMR (151MHz, DMSO-d6) δ157.10,143.44,143.05,138.31,131.32,126.26,113.90,105.98,103.64,77.37,77. 17,73.95,70.56,66.67,61.56,55.67,52.19,52.03,47.82,44.50,29.53,24.44,23.20.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 33 N4O7:477.2349; Found:477.2353.

[0192] (21) Preparation of (2R,3S,4S,5R,6R)-2-(hydroxymethyl)-6-(2-(9-methoxy-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)tetrahydro-2H-pyran-3,4,5-triol (IV-18)

[0193]

[0194] The preparation method was the same as that of Example 6-(4). A white solid (IV-18) was obtained by column chromatography, 100 mg, yield 24.2%, melting point 182.9-185.4°C. 1 H NMR (600MHz, DMSO-d6) δ7.32(d,J=8.8Hz,1H),6.87(d,J=2.8Hz,1H),6.80(dd,J=8.8,2.8Hz,1H),4.89(t,J=4.4Hz,1H) ,4.70(dd,J=5.4,2.2Hz,1H),4.60-4.55(m,1H),4.55-4.51(m,1H),4.35(dd,J=4.6,2.9Hz,1H),4.13(d,J=7.4Hz,1H),4 .03-3.95(m,1H),3.93-3.83(m,1H),3.78(s,3H),3.80-3.78(m,1H),3.77-3.73(m,1H),3.73-3.68(m,1H),3.64-3.61(m ,1H),3.54-3.41(m,3H),3.33-3.24(m,4H),2.64-2.57(m,1H),1.83-1.74(m,3H),1.49-1.38(m,2H),1.27-1.19(m,1H). 13 C NMR (151MHz, DMSO-d6) δ157.04,143.47,143.11,138.11,131.18,126.18,113.83,105.88,103.97,75.40,73. 55,70.88,68.45,66.56,60.75,55.65,52.10,51.98,47.87,44.55,29.46,24.31,23.07.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 33 N4O7:477.2349; Found:477.2351.

[0195] (22) Preparation of (2R,3R,4S,5R,6R)-2-(hydroxymethyl)-6-(3-(9-methoxy-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)tetrahydro-2H-pyran-3,4,5-triol (IV-19)

[0196]

[0197] The preparation method was the same as that of Example 6-(4). A yellow solid (IV-19) was obtained by column chromatography, 140 mg, with a yield of 21.4% and a melting point of 101.8-104.7°C. 1 H NMR (600MHz, DMSO-d6) δ7.31(d,J=8.8Hz,1H),6.87(d,J=2.8Hz,1H),6.79(dd,J=8.8,2.8Hz,1H),5.01(dd,J=12.6,4.9Hz,1H),4.93(dd, J=4.9,1.0Hz,1H),4.89(d,J=5.2Hz,1H),4.61-4.57(m,1H),4.46(t,J=5.9Hz,1H),4.16(dd,J=7.8,3.4Hz,1H),3.88-3.83(m,1H),3.78( s,3H),3.72-3.65(m,3H),3.61-3.56(m,1H),3.55-3.50(m,1H),3.48-3.40(m,2H),3.32-3.28(m,1H),3.17-3.13(m,1H),3.12-3.08(m,1 H),3.07-3.02(m,1H),3.01-2.95(m,1H),2.65-2.59(m,1H),1.97-1.86(m,2H),1.82-1.76(m,3H),1.47-1.42(m,2H),1.27-1.20(m,1H). 13 C NMR(151MHz,DMSO-d6)δ157.06,143.58,143.02,138.35,131.21,126.19,113.79,105.93,103.17,77.17,76.90, 73.79,70.44,66.80,61.45,55.67,52.00,51.33,45.19,44.50,29.60,26.81,24.35,23.18.HRMS(ESI):m / z[M+H] + Calcd for C 24 H 35 N4O7:491.2506; Found:491.2508.

[0198] (23) Preparation of (2R,3S,4S,5R,6R)-2-(hydroxymethyl)-6-(3-(9-methoxy-1,2,3,4,4a,5-hexahydro-6H-pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)tetrahydro-2H-pyran-3,4,5-triol (IV-20)

[0199]

[0200] The preparation method was the same as that of Example 6-(4). A yellow solid (IV-20) was obtained by column chromatography, 130 mg, with a yield of 21.8% and a melting point of 69.8-72.4°C. 1 H NMR (600MHz, DMSO-d6) δ7.31(d,J=8.8Hz,1H),6.87(d,J=2.8Hz,1H),6.79(dd,J=8.8,2.8Hz,1H),4.86(dd,J=12.0,4.6Hz ,1H),4.70(d,J=5.4Hz,1H),4.61-4.56(m,1H),4.56-4.53(m,1H),4.34(dd,J=4.6,2.5Hz,1H),4.11(dd,J=7.4,3.8Hz,1H ),3.85v3.81(m,1H),3.78(s,3H),3.74-3.62(m,3H),3.61-3.56(m,1H),3.56-3.50(m,2H),3.50-3.45(m,1H),3.37-3.35 (m,1H),3.34-3.26(m,4H),2.64-2.58(m,1H),1.97-1.86(m,2H),1.83-1.75(m,3H),1.49-1.40(m,2H),1.27-1.20(m,1H). 13 C NMR(151MHz,DMSO-d6)δ157.08,143.57,142.98,138.45,131.27,126.22,113.81,105.98,103.81,75.64,73.94, 71.10,68.60,66.67,60.91,55.67,52.04,51.42,45.23,44.45,29.56,26.95,24.41,23.23.HRMS(ESI):m / z[M+H] + Calcd for C 24 H 35 N4O7:491.2506; Found:491.2505.

[0201] (24) Preparation of (2R,3R,4S,5R,6R)-2-(2-(9-chloro-1,2,3,4,4a,5-6H-hexahydro[1',2':1,6]pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-21)

[0202]

[0203] The preparation method was the same as that of Example 6-(4). A white solid (IV-21) was obtained by column chromatography, 110 mg, with a yield of 29.7% and a melting point of 130.7-106.6°C. 1 H NMR (600MHz, DMSO-d6) δ7.38(d,J=8.6Hz,1H),7.36(d,J=2.4Hz,1H),7.15(dd,J=8.6,2.4Hz,1H),5.03(dd,J=5.1,3.8Hz,1H),4.93(dd,J= 4.9,3.9Hz,1H),4.89(dd,J=5.3,1.2Hz,1H),4.67-4.62(m,1H),4.48-4.42(m,1H),4.19(dd,J=7.8,1.3Hz,1H),4.03-3.96(m,1H),3.94-3. 86(m,1H),3.83-3.76(m,1H),3.76-3.73(m,1H),3.73-3.68(m,1H),3 .67-3.63(m,1H),3.46-3.39(m,2H),3.39-3.36(m,1H),3.15-3.11(m ,1H),3.11-3.07(m,1H),3.07-3.01(m,1H),3.00-2.93(m,1H),2.71- 2.65(m,1H),1.85-1.75(m,3H),1.50-1.40(m,2H),1.28-1.21(m,1H). 13 C NMR(151MHz,DMSO-d6)δ144.44,143.68,137.86,135.35,128.40,126.57,123.94,123.42,103.27,76.89, 76.65,73.67,70.30,66.49,61.30,51.95,51.70,47.94,44.46,29.48,24.23,22.94.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 30 N4O6Cl:481.1854; Found:481.1856.

[0204] (25) Preparation of (2R,3S,4S,5R,6R)-2-(2-(9-chloro-1,2,3,4,4a,5-hexahydro-6H-hexahydro[1',2':1,6]pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)ethoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-22)

[0205]

[0206] The preparation method was the same as that of Example 6-(4). A pale yellow solid (IV-22) was obtained by column chromatography, 120 mg, with a yield of 27.3% and a melting point of 202.2-205.0°C. 1 H NMR (600MHz, DMSO-d6) δ7.38(d,J=8.7Hz,1H),7.36(d,J=2.4Hz,1H),7.14(dd,J=8.6,2.5Hz,1H),4.90-4.85(m,1 H),4.70(dd,J=5.4,2.1Hz,1H),4.67-4.62(m,1H),4.54-4.51(m,1H),4.36-4.33(m,1H),4.13(d,J=7.3Hz,1H),4 .01-3.95(m,1H),3.91-3.84(m,1H),3.80-3.66(m,3H),3.63-3.61(m,1H),3.53-3.40(m,3H),3.40-3.35(m,1H), 3.33-3.27(m,2H),3.27-3.24(m,1H),2.70-2.64(m,1H),1.83-1.76(m,3H),1.50-1.38(m,2H),1.28-1.21(m,1H). 13 C NMR(151MHz,DMSO-d6)δ144.45,143.66,137.96,135.42,128.34,126.63,123.87,123.47,103.97,75.40, 73.54,70.88,68.42,66.32,60.71,51.94,51.77,48.06,44.43,29.48,24.27,22.99.HRMS(ESI):m / z[M+H] + Calcd for C 22 H 29 N4O6ClNa:503.1673; Found:503.1670.

[0207] (26) Preparation of (2R,3R,4S,5R,6R)-2-(3-(9-chloro-1,2,3,4,4a,5-6H-hexahydro[1',2':1,6]pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-23)

[0208]

[0209] The preparation method was the same as that of Example 6-(4). A light yellow solid (IV-23) was obtained by column chromatography, 100 mg, yield 13.0%, melting point 118.8-121.5°C. 1 H NMR (600MHz, DMSO-d6) δ7.37(d,J=3.9Hz,1H),7.36(d,J=2.2Hz,1H),,7.13(dd,J=8.6,2.4Hz,1H),5.01(dd,J=13.7,4.9Hz,1H), 4.93(dd,J=4.9,1.2Hz,1H),4.89(d,J=5.2Hz,1H),4.67-4.63(m,1H),4.46(t,J=5.9Hz,1H),4.15(dd,J=7.8,3.9Hz,1H),3.88-3. 82(m,1H),3.73-3.60(m,4H),3.55-3.50(m,1H),3.46-3.42(m,1H),3.42-3.35(m,2H),3.17-3.13(m,1H),3.11-3.08(m,1H),3.0 7-3.02(m,1H),3.00-2.95(m,1H),2.71-2.65(m,1H),1.98-1.85(m,2H),1.84-1.75(m,3H),1.51-1.40(m,2H),1.29-1.22(m,1H). 13 C NMR(151MHz,DMSO-d6)δ144.38,143.77,138.09,135.36,128.32,126.58,123.80,123.48,103.05,76.97,76.76, 73.76,61.36,66.62,61.36,51.94,50.89,45.36,44.46,39.02,29.58,26.66,24.24,23.00.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 32 N4O6Cl:495.2010; Found:495.2012.

[0210] (27) Preparation of (2R,3S,4S,5R,6R)-2-(3-(9-chloro-1,2,3,4,4a,5-6H-hexahydro[1',2':1,6]pyrido[1',2':1,6]pyrazino[2,3-b]quinoxalin-6-yl)propoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (IV-24)

[0211]

[0212] The preparation method was the same as that of Example 6-(4). A white solid (IV-24) was obtained by column chromatography, 80 mg, with a yield of 11.9% and a melting point of 83.5-86.3°C. 1 H NMR (600MHz, DMSO-d6) δ7.38-7.37(m,1H),7.37-7.36(m,1H),7.13(dd,J=8.6,2.4Hz,1H),4.85(dd,J=13 .0,4.6Hz,1H),4.69(d,J=5.4Hz,1H),4.67-4.63(m,1H),4.55-4.52(m,1H),4.35-4.33(m,1H),4.10(dd, J=7.4,4.4Hz,1H),3.85-3.80(m,1H),3.75-3.60(m,4H),3.56-3.45(m,3H),3.42-3.35(m,2H),3.33-3.2 6(m,3H),2.71-2.65(m,1H),1.97-1.86(m,2H),1.84-1.76(m,3H),1.51-1.40(m,2H),1.29-1.21(m,1H). 13 CNMR(151MHz,DMSO-d6)δ144.39,143.79,138.29,135.51,128.26,126.66,123.70,123.65,103.80,75.66,73. 93,71.10,68.60,66.29,60.90,51.94,51.11,45.41,,45.39,29.61,26.86,24.36,23.12.HRMS(ESI):m / z[M+H] + Calcd for C 23 H 32 N4O6Cl:495.2010; Found:495.2015.

[0213] In the third aspect, the beneficial effects of the present invention are demonstrated by specific experimental examples as follows;

[0214] Experimental Example 1: CCK-8 method to detect the anti-tumor activity of anticancer drug TNBG-sugar conjugate

[0215] The CCK-8 (cell counting kit-8) assay was used to test the antitumor activity of the TNBG-sugar conjugates shown in Formula I and Formula IV against the human lung adenocarcinoma cell line A549, with TNBG and sorafenib as positive controls. A549 cells were inoculated in DMEM medium containing 10% FBS (fetal bovine serum) and cultured in a cell incubator at 37°C and 5% CO2. The cells were centrifuged and resuspended in the logarithmic growth phase, then planted in a 96-well plate. After the cells had fully adhered to the wall and grown, the specified concentration of TNBG-sugar conjugate was added to the well plate and incubated for 72 hours. The original culture medium was discarded, 10% CCK-8 working solution was added, and the OD value was measured at a wavelength of 450nm after incubation in the incubator for 40 minutes. SPSS21.0 software was used for data analysis, with the inhibition rate as the response frequency and the drug concentration as the dependent variable. The concentration was Logit transformed, the observed values ​​were summarized as a total of 100, and probit regression analysis was performed to calculate the IC 50 The final test result is IC 50 The antitumor activity of the anticancer drug TNBG-sugar conjugate of the present invention on A549 cells is shown in Table 1.

[0216] Table 1 Antitumor activity of anticancer drug TNBG-sugar conjugate on A549 cell line

[0217]

[0218] Note: “a” represents the average value of three repeated experiments.

[0219] As can be seen from Table 1, the anticancer drug TNBG-sugar conjugates IV-12, IV-14 and IV-15 all showed good anti-tumor activity and had good application prospects.

Claims

1. A compound of formula I or a pharmaceutically acceptable salt or a stereoisomer thereof, in, n is an integer selected from 1 to 6; G is selected from monosaccharides (glucose, galactose, mannose, etc.), polysaccharides (dextran, lactose, etc.), etc.; R1-R8 is selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxyl, nitro, amino, carboxyl, formula II or formula III, etc.; n1 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond; R9 and R 10 Selected from hydrogen, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl; and methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl having a substituent; n2 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond; n3 is an integer selected from 1 to 2, the symbol represents a single bond or a double bond; X is selected from oxygen, nitrogen or carbon; R 11 Selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxyl, nitro, amino or carboxyl, etc.

2. The compound of formula I according to claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein: n is often selected from the integer 3, and G can be selected from D-glucose, D-galactose, and the like.

3. The synthesis of the anticancer drug TNBG-sugar conjugate shown in Formula I has the following characteristics: First, the synthesis was performed according to the mature route of the research group to obtain the lead compound of the anticancer drug TNBG, i.e., the key intermediate 10, and then the target compound of formula I was obtained through substitution reaction, glycosylation reaction, and deacetylation reaction; The substitution reaction is that the hydrogen on the secondary amine is replaced by 1-bromoethanol or propanol under strong alkaline conditions (NaH (60%)); the glycosidation reaction is that the hydroxyl group reacts with the chirally fixed tetraacetyl brominated sugar under the catalysis of heavy metal salts; the deacetylation reaction is carried out in a methanol solution in an alkaline (potassium carbonate) environment to obtain the target compounds I-1 and I-2.

4. Another compound of formula IV or a pharmaceutically acceptable salt or a stereoisomer thereof, in, n is an integer selected from 1 to 6; G is selected from monosaccharides (glucose, galactose, mannose, etc.), polysaccharides (dextran, lactose, etc.), etc.; R1-R4 is selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxyl, nitro, amino, carboxyl, formula V or formula VI, etc.; n1 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond; R5 and R6 are selected from hydrogen, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl; and methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, cyclopropyl or cyclobutyl having a substituent; n2 is an integer selected from 1 to 8, the symbol represents a single bond or a double bond; n3 is an integer selected from 1 to 2, the symbol represents a single bond or a double bond; X is selected from oxygen, nitrogen or carbon; R7 is selected from hydrogen, halogen, C1-C8 alkyl, C1-C8 alkoxy, hydroxy, nitro, amino or carboxyl, etc.

5. The compound of formula IV according to claim 4, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein: n is often selected from integers 2 and 3; R 1-4 wherein is hydrogen, alkyl is preferably methyl, halogen is preferably F, Cl, alkoxy is preferably methoxy, etc.; G is preferably D-glucose, D-galactose, etc.

6. The synthesis of the anticancer drug TNBG-sugar conjugate shown in Formula IV has the following characteristics: The synthesis is carried out through cyclization reaction, chlorination reaction, and cyclization reaction to obtain the key intermediate 14, and then through substitution reaction, glycosylation reaction, and deacetylation reaction to obtain the target compound of formula IV; The cyclization reaction is that the starting material reacts with diethyl oxalate under acidic conditions (dilute hydrochloric acid); the chlorination reaction is that phosphorus oxychloride is used to convert the hydroxyl group into a chlorine atom; the cyclization reaction is that under alkaline conditions (potassium carbonate) the cyclization reaction is carried out with 2-aminomethylpiperidine to form a four-membered ring key intermediate 14; the substitution reaction is that the hydrogen on the secondary amine is replaced by 1-bromoethanol or propanol under strong alkaline conditions (NaH (60%)); the glycosidation reaction is that the hydroxyl group reacts with a chirally fixed tetraacetyl brominated sugar under the catalysis of a heavy metal salt; the deacetylation reaction is carried out in a methanol solution in an alkaline environment (potassium carbonate) to obtain the target compounds IV-1 to IV-24.

7. The compound represented by formula I and IV or a pharmaceutically acceptable salt or a stereoisomer thereof, characterized in that: The compounds described herein are representative compounds such as the following:

8. Use of the compound according to claim 7 or a pharmaceutically acceptable salt thereof or a stereoisomer thereof in the preparation of a drug useful for treating and / or preventing cancer.

9. The use according to claim 8, characterized in that: The prepared anticancer drug is a drug for treating cancer.

10. The use according to claim 9, characterized in that: Preferably, the cancer is lung cancer, liver cancer, etc.

11. A medicine, which is a preparation prepared by using the compound according to claim 7 or its pharmaceutically acceptable salt or stereoisomer thereof as an active ingredient and adding suitable excipients or auxiliary ingredients.

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