Synthesis method of trivalent GalNAc compound

Abstract

The invention discloses a synthesis method of a trivalent GalNAc compound, which comprises the following steps: under the action of a condensing agent and organic alkali, reacting carboxyl-containing D-galactosamine pentaacetate as shown in a formula I with carbobenzoxy-protected diamine as shown in a formula II to obtain an intermediate III; under the hydrogen atmosphere, the intermediate III reacts under the catalysis of palladium carbon to remove a carbobenzoxy protecting group, and an intermediate IV is obtained; under the action of a condensing agent and organic alkali, the intermediate IV reacts with tri [(2-carboxyl ethyoxyl) methyl] methyl benzyl carbamate, and the trivalent GalNAc compound shown in the formula V is obtained. The method is simple and rapid in reaction process, mild in condition, simple and convenient in post-treatment, and high in target product yield which at least reaches about 55% and is obviously higher than that in the prior art.

Description

Technical Field

[0001] The invention belongs to the field of organic synthesis and relates to a method for synthesizing trivalent GalNAc compounds. Background Art

[0002] N-acetylgalactosamine (GalNAc) is a targeted ligand for the asialoprotein receptor (ASGPR) overexpressed by hepatocytes and has many applications in the biomedical field. Compared with traditional monovalent GalNAc, trivalent GalNAc can specifically bind to the sugar recognition domains of multiple subunits of ASGPR, showing stronger affinity and specificity for ASGPR. Givosiran, which was launched in 2020, is a polysaccharide-small nucleic acid conjugate. With the binding ability between trivalent GalNAc and ASGPR specifically expressed by hepatocytes, the drug is accurately delivered to the liver, and the drug enters the cell to function through endocytosis, which is used to treat acute hepatic porphyria. In addition, lysosome-targeted chimeras are a type of drug that has attracted much attention in recent years. This technology also uses trivalent GalNAc to mediate the degradation of hepatocyte-related proteins. Therefore, trivalent GalNAc has great prospects for clinical application.

[0003] The synthesis process of trivalent GalNAc compounds is complicated. The representative method is to first condense propylene diamine with a tricarboxylic acid protected by a benzyloxycarbonyl group, and then couple it with a galactose unit to obtain the target molecule. The typical synthesis route is as follows:

[0004]

[0005] The above three steps are the rate-limiting steps in the existing trivalent GalNAc synthesis route, and have a low yield, with the total yield of the three steps being only 32%. The purification steps are relatively complicated, and two steps require column chromatography purification, resulting in the problem of excessively high costs. Summary of the invention

[0006] The purpose of the present invention is to provide a method for synthesizing trivalent GalNAc compounds with a simple reaction process, mild conditions, high yield and convenient post-processing, in view of the shortcomings of the prior art such as low yield and complicated purification process.

[0007] In order to achieve the above object, the present invention adopts the following technical solution:

[0008] A method for synthesizing a trivalent GalNAc compound, the synthesis route is as follows:

[0009]

[0010] Wherein, m is a positive integer from 1 to 10, and n is a positive integer from 1 to 10;

[0011] The steps include:

[0012] Step (1), using one of dichloromethane, acetonitrile or tetrahydrofuran as a reaction solvent, in the presence of a condensing agent and an organic base, the carboxyl-containing D-aminogalactose pentaacetate of formula I and the benzyloxycarbonyl-protected diamine of formula II react to obtain intermediate III;

[0013] Step (2), using tetrahydrofuran or C1-C4 lower alcohol as the reaction solvent, in a hydrogen atmosphere, intermediate III is catalyzed by palladium carbon to remove the benzyloxycarbonyl protecting group to obtain intermediate IV;

[0014] Step (3), using at least one of N,N-dimethylformamide, dimethyl sulfoxide or tetrahydrofuran or a mixed solvent selected from one of N,N-dimethylformamide or dimethyl sulfoxide and one selected from dichloromethane or acetonitrile as the reaction solvent, in the presence of a condensing agent and an organic base, the intermediate IV reacts with benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate (compound 5) to obtain a trivalent GalNAc compound with a structure as shown in Formula V.

[0015] Specifically, m is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, and n is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0016] In some embodiments, m=3, n=2; m=3, n=1; m=3, n=4; m=3, n=6; m=3, n=8; m=5, n=2; m=7, n=2.

[0017] In step (1), preferably, the reaction solvent is dichloromethane or acetonitrile.

[0018] The molar ratio of the carboxyl-containing D-galactosamine pentaacetate to the benzyloxycarbonyl-protected diamine is 1:1 to 1:1.2, preferably 1:1.04 to 1:1.06.

[0019] The feed concentration of the carboxyl-containing D-galactosamine pentaacetate (i.e. the ratio of the molar amount of the carboxyl-containing D-galactosamine pentaacetate to the volume of the reaction solvent) is 0.1-1 mol / L, preferably 0.1-0.3 mol / L.

[0020] The condensing agent is selected from HATU, HBTU or HCTU, preferably HATU or HBTU.

[0021] The organic base is selected from N,N-diisopropylethylamine, triethylamine, 1,8-diazabicycloundec-7-ene or N-methylmorpholine, preferably N,N-diisopropylethylamine, triethylamine or N-methylmorpholine.

[0022] Specifically, the reaction solvent is dichloromethane or acetonitrile, the condensation agent is HATU or HBTU, and the organic base is N,N-diisopropylethylamine or triethylamine; the reaction solvent is dichloromethane or acetonitrile, the condensation agent is HATU, and the organic base is N-methylmorpholine; the reaction solvent is dichloromethane, the condensation agent is HATU, and the organic base is 1,8-diazabicycloundec-7-ene; the reaction solvent is dichloromethane, the condensation agent is HCTU, and the organic base is N,N-diisopropylethylamine or triethylamine; the reaction solvent is tetrahydrofuran, the condensation agent is HATU, and the organic base is N,N-diisopropylethylamine or triethylamine.

[0023] The molar ratio of the carboxyl-containing D-galactosamine pentaacetate to the condensing agent is 1:1.1 to 1:1.5, preferably 1:1.15 to 1:1.25.

[0024] The molar ratio of the carboxyl-containing D-galactosamine pentaacetate to the organic base is 1:1.5 to 1:3.

[0025] The reaction time is no more than 10 minutes, preferably 5 to 10 minutes; the reaction temperature is 10 to 30°C, preferably 20 to 30°C.

[0026] Specifically, the carboxyl-containing D-galactosamine pentaacetate shown in formula I is dissolved in a reaction solvent, a condensing agent and an organic base are added, stirred for 10 to 40 minutes, preferably 30 minutes, and then the benzyloxycarbonyl-protected diamine shown in formula II is added to react to obtain intermediate III.

[0027] After the reaction was completed, ethyl acetate was added to the reaction solution, and the mixture was washed with a saturated aqueous solution of ammonium chloride. The organic phase was taken, dried over anhydrous sodium sulfate, and concentrated to obtain Intermediate III.

[0028] In step (2), preferably, the C1-C4 lower alcohol is methanol.

[0029] The feed concentration of intermediate III is 0.1-1 mol / L.

[0030] Calculated in terms of palladium, the molar ratio of palladium carbon to intermediate III is 0.05:1 to 0.2:1.

[0031] The loading amount of palladium in palladium carbon is 5-10%.

[0032] The hydrogen pressure is 1 to 4 standard atmospheres.

[0033] The reaction time is 4 to 6 hours; the reaction temperature is 10 to 30° C., preferably 20 to 25° C.; the reaction is carried out under stirring.

[0034] Specifically, the intermediate III is dissolved in tetrahydrofuran or methanol and other C1-C4 lower alcohols, palladium carbon is added, and then hydrogen is introduced. The reaction is stirred to remove the benzyloxycarbonyl protecting group from the intermediate III to obtain the intermediate IV.

[0035] After the reaction was completed, the mixture was filtered and the filtrate was concentrated to obtain compound IV.

[0036] In step (3), the condensing agent is selected from HATU or HBTU.

[0037] The organic base is selected from N,N-diisopropylethylamine, triethylamine or N-methylmorpholine.

[0038] Specifically, the reaction solvent is N,N-dimethylformamide, dimethyl sulfoxide, tetrahydrofuran, or a mixed solvent selected from one of N,N-dimethylformamide or dimethyl sulfoxide and one selected from dichloromethane or acetonitrile, the condensing agent is HATU or HBTU, and the organic base is N,N-diisopropylethylamine or triethylamine; the reaction solvent is N,N-dimethylformamide, dimethyl sulfoxide, tetrahydrofuran, or a mixed solvent selected from one of N,N-dimethylformamide or dimethyl sulfoxide and one selected from dichloromethane or acetonitrile, the condensing agent is HATU or HBTU, and the organic base is N-methylmorpholine.

[0039] The volume ratio of one selected from N,N-dimethylformamide or dimethyl sulfoxide to one selected from dichloromethane or acetonitrile is 1:1.

[0040] Considering that N,N-dimethylformamide and dimethyl sulfoxide have good solubility and reaction yield, but have high boiling points, which leads to troublesome post-processing, the reaction solvent is preferably tetrahydrofuran with a lower boiling point.

[0041] The molar ratio of benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate to intermediate IV is 1:3 to 1:4, preferably 1:3.6.

[0042] The molar ratio of benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate to the condensing agent is 1:3.3 to 1:5, preferably 1:3.6.

[0043] The molar ratio of benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate to the organic base is 1:5 to 1:7, preferably 1:6.

[0044] The feed concentration of benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate is 0.1 to 1 mol / L.

[0045] The reaction time is 6 to 8 hours; the reaction temperature is 10 to 30° C., preferably 20 to 25° C.; the reaction is carried out under stirring.

[0046] Specifically, compound 5, i.e. benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate, is dissolved in a reaction solvent, a condensing agent and an organic base are added, stirred for 20 to 50 minutes, preferably for 40 minutes, and amino-containing D-galactosamine pentaacetate shown in intermediate IV is added to react to obtain a trivalent GalNAc compound.

[0047] After the reaction is completed, the reaction solution is concentrated and then subjected to normal phase silica gel column chromatography using a mixed solvent of dichloromethane and methanol in a volume ratio of 20:1 to 4:1 as an eluent to purify and obtain trivalent GalNAc compounds.

[0048] Beneficial technical effects of the present invention:

[0049] The present invention provides a method for synthesizing a trivalent GalNAc compound, the reaction process is simple and rapid, the conditions are mild, and the target product has a high yield, the yield is at least about 55%, which is significantly higher than about 32% in the prior art, and the post-processing is convenient. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1 This is the hydrogen nuclear magnetic resonance spectrum of the trivalent GalNAc compound obtained in Example 1. DETAILED DESCRIPTION

[0051] In order to further illustrate the technical solution of the present invention, the following examples are given, and comparative examples are given to compare with the preparation method provided by the present invention. The following examples are purely illustrative and are only used to specifically describe the technical solution of the present invention, and cannot be understood as limiting the scope of protection of the present invention.

[0052] DCM is dichloromethane, MeCN is acetonitrile, THF is tetrahydrofuran, DMF is N,N-dimethylformamide, DIPEA is N,N-diisopropylethylamine, TEA is triethylamine, DBU is 1,8-diazabicycloundec-7-ene, NMM is N-methylmorpholine, and Py is pyridine.

[0053] The room temperature is 25℃.

[0054] Example 1

[0055]

[0056] Compound 1 (2.20 g, 4.92 mmol) was dissolved in dichloromethane (40 mL), and HATU (2.24 g, 5.91 mmol) and N,N-diisopropylethylamine (1.71 mL, 9.84 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 2 (1.08 g, 5.17 mmol) was added, and the mixture was stirred at room temperature for 5 min. After the reaction, the reaction solution was diluted with ethyl acetate (200 mL), and washed three times with saturated aqueous ammonium chloride solution (100 mL×3), and the organic phase was dried over anhydrous sodium sulfate and concentrated to obtain compound 3 (2.89 g, yield 92%).

[0057] 1 H NMR (300MHz, DMSO-d6) δ7.84 (d, J=9.2Hz, 1H, NH), 7.81-7.67 (m, 1H, NH), 7.40-7.30 (m, 5H, ArH), 7.29-7 .18(m,1H,NH),5.22(d,J=3.4Hz,1H,CH),5.07-4.91(m,3H,CH,CH2),4.48(d,J=8.5Hz,1H,CH),4.03(s,2 H,CH2),3.94-3.82(m,1H,CH),3.76-3.65(m,1H,CH),3.47-3.37(m,2H,CH2),3.10-2.93(m,4H,2CH2),2. 11(s,3H,CH3),2.06-1.95(m,5H,CH2,CH3),1.90(s,3H,CH3),1.77(s,3H,CH3),1.61-1.34(m,6H,3CH2).

[0058]

[0059] Compound 3 (0.98 g, 1.54 mmol) was dissolved in methanol (13 mL), palladium carbon (10%, 0.17 g, 0.16 mmol) was added, and then hydrogen was introduced, maintaining the hydrogen pressure at 1 to 4 standard atmospheres. The reaction was stirred at room temperature for 5 h. After the reaction, the reaction solution was filtered through diatomaceous earth, and the filtrate was concentrated to obtain compound 4 (0.96 g, yield 98%).

[0060]

[0061] Compound 5 (0.65 g, 1.38 mmol) was dissolved in tetrahydrofuran (10 mL), HATU (1.89 g, 4.97 mmol) and N,N-diisopropylethylamine (1.44 mL, 8.28 mmol) were added, and the mixture was stirred at room temperature for 40 min. Compound 4 (2.51 g, 4.97 mmol) was added, and the mixture was stirred at room temperature for 8 h. After the reaction was completed, the reaction solution was concentrated, and the crude product was subjected to normal phase silica gel column chromatography with dichloromethane / methanol (4:1 v / v) as the eluent, and the trivalent GalNAc compound (1.97 g, 73%) with the structure shown in Formula V1 was obtained by separation and purification.

[0062] The H NMR spectra of trivalent GalNAc compounds are shown in Figure 1 . 1 H NMR (300MHz, DMSO-d6) δ7.95-7.82(m,6H,6NH),7.80-7.71(m,3H,3NH),7.37-7.31(m,5H,5ArH),6.58(s,1H,NH),5.21(d,J= 3.4Hz,3H,3CH),5.02-4.93(m,5H,CH2,3CH),4.49(d,J=8.3Hz,3H,3CH),4.03(s,6H,3CH2),3.95-3.80(m,3H,3CH),3.77-3.6 5(m,3H,3CH),3.58-3.52(m,6H,3CH2),3.48(s,6H,3CH2),3.44-3.39(m,6H,3CH2),3.12-2.93(m,12H,6CH2),2.32-2.24(m,6 H,3CH2),2.10(s,9H,3CH3),2.07-1.96(m,15H,3CH2,3CH3),1.89(s,9H,3CH3),1.78(s,9H,3CH3),1.55-1.40(m,18H,6CH3).

[0063] The total yield of the three steps in this embodiment is 66%, which is much higher than the existing synthesis method (32%).

[0064] Comparative Example 1

[0065] The present invention uses benzyloxycarbonyl protected diamine as raw material. Similarly, tert-butyloxycarbonyl protected diamine is also a common raw material for preparing trivalent GalNAc.

[0066] Using tert-butyloxycarbonyl-protected propylenediamine (compound 6) instead of benzyloxycarbonyl-protected propylenediamine (compound 2) as a raw material to prepare trivalent GalNAc, the steps are as follows:

[0067]

[0068] Compound 1 (0.57 g, 1.27 mmol) was dissolved in dichloromethane (10 mL), HATU (0.58 g, 1.52 mmol) and N,N-diisopropylethylamine (0.44 mL, 2.54 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 6 (0.23 mg, 1.33 mmol) was then added and stirred at room temperature for 10 min. After the reaction was completed, the reaction solution was diluted with ethyl acetate (50 mL) and washed three times with saturated aqueous ammonium chloride solution (25 mL×3). The organic phase was dried over anhydrous sodium sulfate and concentrated to obtain compound 7 (0.52 g, yield 68%).

[0069]

[0070] Compound 7 (0.33 g, 0.55 mmol) was dissolved in dichloromethane (5 mL), trifluoroacetic acid (1 mL) was slowly added dropwise, and stirred at room temperature for 1 h. After the reaction was completed, the reaction solution was concentrated to obtain compound 8 (0.30 g, yield 97%).

[0071]

[0072] Compound 5 (0.33 g, 0.70 mmol) was dissolved in tetrahydrofuran (6 mL), HATU (0.96 g, 2.52 mmol) and N,N-diisopropylethylamine (0.73 mL, 4.20 mmol) were added, and the mixture was stirred at room temperature for 40 min. Compound 8 (1.55 g, 2.52 mmol) was added, and the mixture was stirred at room temperature for 10 h. After the reaction was completed, the reaction solution was concentrated, and the crude product was subjected to normal phase silica gel column chromatography with dichloromethane / methanol (4:1 v / v) as the eluent, and trivalent GalNAc compounds (0.79 g, 59%) were obtained by separation and purification.

[0073] In the first step of post-treatment in Comparative Example 1, repeated washing with a saturated aqueous solution of ammonium chloride is required to remove impurities such as organic bases, but since compound 7 has a certain water solubility, part of the product is washed away by water; the amino group used in the third step is trifluoroacetate, and a condensation product of trifluoroacetic acid and sugar exists in the condensation process, so the total yield of the three steps is only 39%, which is lower than the total yield (66%) of Example 1 of the present invention.

[0074] In addition, since the tert-butyloxycarbonyl group can only be removed by acid hydrolysis, and the trifluoroacetic acid used in the second step has a high boiling point, rotary evaporation is difficult, and a large amount of acidic gas will damage the pump.

[0075] Example 2

[0076] Using compound 1 (m=3) and compound 2 (n=2) as raw materials, the effect of reaction conditions on the yield was investigated.

[0077] According to the feed concentration of compound 1 being 0.3 mol / L, compound 1 (1 mmol) was dissolved in the reaction solvent, and a condensing agent (1.2 mmol) and an organic base (2 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 2 (1.05 mmol) was then added, and the mixture was stirred at room temperature for 10 min. The reaction solution was diluted with ethyl acetate and washed three times with a saturated aqueous ammonium chloride solution. The volume of the saturated aqueous ammonium chloride solution for each washing was 0.5 times the volume of ethyl acetate. The organic phase was then dried over anhydrous sodium sulfate and concentrated to obtain compound 3.

[0078] Table 1. Effect of reaction conditions on yield

[0079]

[0080] Example 3

[0081] Using compound 4 (m=3, n=2) and compound 5 as raw materials, the effect of reaction conditions on the yield was investigated.

[0082] According to the feed concentration of compound 5 being 0.1 mol / L, compound 5 was dissolved in the reaction solvent, a condensing agent and an organic base were added, and the mixture was stirred at room temperature for 40 min. Compound 4 (2.51 g, 4.97 mmol) was added, and the mixture was stirred at room temperature for 8 h. After the reaction, the reaction solution was concentrated, and the crude product was subjected to normal phase silica gel column chromatography with dichloromethane / methanol (4:1 v / v) as the eluent, and the trivalent GalNAc compound with the structure shown in Formula V1 was obtained by separation and purification.

[0083] Among them, the molar ratio of compound 4 to compound 5 is 3.6:1; the molar ratio of the condensing agent to compound 5 is 3.6:1; and the molar ratio of the organic base to compound 5 is 6:1.

[0084] Table 2. Effect of reaction conditions on the yield of trivalent GalNAc compounds a

[0085]

[0086]

[0087] Example 4

[0088]

[0089] Compound 1 (0.87 g, 1.95 mmol) was dissolved in dichloromethane (10 mL), HATU (0.89 g, 2.34 mmol) and N,N-diisopropylethylamine (0.68 mL, 3.90 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 9 (0.40 g, 2.05 mmol) was then added and stirred at room temperature for 10 min. After the reaction was completed, the reaction solution was diluted with ethyl acetate (50 mL), washed three times with saturated aqueous ammonium chloride solution (25 mL×3), dried over anhydrous sodium sulfate, and concentrated to obtain compound 10 (1.13 g, yield 93%).

[0090] 1 H NMR (300MHz, DMSO-d6) δ7.92 (d, J=9.3Hz, 1H, NH), 7.84-7.63 (m, 1H, NH), 7.44-7.29 (m, 5H, ArH), 7.28-7 .17(m,1H,NH),5.23(d,J=4.2Hz,1H,CH),5.23-4.95(m,3H,CH,CH2),4.49(d,J=8.6Hz,1H,CH),4.10(s,2 H,CH2),3.97-3.80(m,1H,CH),3.80-3.66(m,1H,CH),3.49-3.38(m,2H,CH2),3.20-2.94(m,4H,2CH2),2. 14(s,3H,CH3),2.10-1.96(m,5H,CH2,CH3),1.95(s,3H,CH3),1.82(s,3H,CH3),1.52-1.38(m,4H,2CH2).

[0091] Example 5

[0092]

[0093] Compound 1 (0.75 g, 1.67 mmol) was dissolved in dichloromethane (10 mL), HATU (0.76 g, 2.00 mmol) and N,N-diisopropylethylamine (0.58 mL, 3.34 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 11 (0.41 mg, 1.75 mmol) was added and stirred at room temperature for 10 min. After the reaction, the reaction solution was diluted with ethyl acetate (50 mL), washed three times with saturated aqueous ammonium chloride solution (25 mL×3), dried over anhydrous sodium sulfate, and concentrated to obtain compound 12 (1.01 g, yield 90%).

[0094] 1H NMR (300MHz, DMSO-d6) δ7.93 (d, J=9.2Hz, 1H, NH), 7.87-7.69 (m, 1H, NH), 7.46-7.26 (m, 5H, ArH), 7.31-7.13(m,1H,NH),5.31(d,J=3.9Hz,1H,CH),5.26-4.97(m,3H,CH,CH2),4.54(d,J=8.8Hz,1H, CH),4.12(s,2H,CH2),4.00-3.81(m,1H,CH),3.75-3.62(m,1H,CH),3.51-3.38(m,2H,CH2),3.22- 2.91(m,4H,2CH2),2.20(s,3H,CH3),2.12-1.99(m,5H,CH2,CH3),1.90-1.29(m,16H,2CH3,5CH2).

[0095] Example 6

[0096]

[0097] Compound 1 (0.59 g, 1.32 mmol) was dissolved in dichloromethane (10 mL), HATU (0.60 g, 1.58 mmol) and N,N-diisopropylethylamine (0.46 mL, 2.64 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 13 (0.36 mg, 1.38 mmol) was added and stirred at room temperature for 10 min. After the reaction, the reaction solution was diluted with ethyl acetate (50 mL), washed three times with saturated aqueous ammonium chloride solution (25 mL×3), dried over anhydrous sodium sulfate, and concentrated to obtain compound 14 (0.80 g, yield 86%).

[0098] 1H NMR(300MHz,DMSO-d6)δ8.01(d,J=9.3Hz,1H,NH),7.93-7.77(m,1H,NH),7.55-7.40(m,5H,ArH), 7.36-7.19(m,1H,NH),5.46(d,J=3.6Hz,1H,CH),5.31-5.03(m,3H,CH,CH2),4.70(d,J=8.6Hz,1H, CH),4.31(s,2H,CH2),4.11-3.93(m,1H,CH),3.83-3.69(m,1H,CH),3.60-3.49(m,2H,CH2),3.30- 2.94(m,4H,2CH2),2.31(s,3H,CH3),2.19-2.01(m,5H,CH2,CH3),1.92-1.09(m,20H,2CH3,7CH2).

[0099] Example 7

[0100]

[0101] Compound 1 (0.59 g, 1.23 mmol) was dissolved in dichloromethane (10 mL), HATU (0.56 g, 1.48 mmol) and N,N-diisopropylethylamine (0.43 mL, 2.46 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 15 (0.38 mg, 1.29 mmol) was added and stirred at room temperature for 10 min. After the reaction, the reaction solution was diluted with ethyl acetate (50 mL), washed three times with saturated aqueous ammonium chloride solution (25 mL×3), dried over anhydrous sodium sulfate, and concentrated to obtain compound 16 (0.78 g, yield 88%).

[0102] 1 H NMR(300MHz,DMSO-d6)δ8.02(d,J=9.7Hz,1H,NH),7.90-7.70(m,1H,NH),7.60-7.42(m,5H, ArH),7.30-7.11(m,1H,NH),5.40(d,J=3.3Hz,1H,CH),5.28-5.07(m,3H,CH,CH2),4.79(d, J=8.9Hz,1H,CH),4.38(s,2H,CH2),4.17-3.89(m,1H,CH),3.81-3.64(m,1H,CH),3.55-3.4 1(m,2H,CH2),3.25-2.82(m,4H,2CH2),2.38(s,3H,CH3),2.21-1.03(m,29H,3CH3,10CH2).

[0103] Example 8

[0104]

[0105] Compound 17 (0.67 g, 1.41 mmol) was dissolved in dichloromethane (10 mL), HATU (0.64 g, 1.69 mmol) and N,N-diisopropylethylamine (0.49 mL, 2.82 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 2 (0.31 mg, 1.48 mmol) was then added and stirred at room temperature for 10 min. After the reaction was completed, the reaction solution was diluted with ethyl acetate (50 mL), washed three times with saturated aqueous ammonium chloride solution (25 mL×3), dried over anhydrous sodium sulfate, and concentrated to obtain compound 18 (0.85 g, yield 90%).

[0106] 1 H NMR(300MHz,DMSO-d6)δ7.92(d,J=9.1Hz,1H,NH),7.82-7.60(m,1H,NH),7.50-7.32(m,5H,ArH), 7.22-7.12(m,1H,NH),5.16(d,J=3.5Hz,1H,CH),4.99-4.86(m,3H,CH,CH2),4.52(d,J=8.1Hz,1H, CH),4.18(s,2H,CH2),3.88-3.75(m,1H,CH),3.69-3.59(m,1H,CH),3.40-3.29(m,2H,CH2),3.18- 2.99(m,4H,2CH2),2.20(s,3H,CH3),2.02-1.90(m,5H,CH2,CH3),1.82-1.42(m,16H,5CH2,2CH3).

[0107] Example 9

[0108]

[0109] Compound 19 (0.76 g, 1.51 mmol) was dissolved in dichloromethane (10 mL), HATU (0.69 g, 1.81 mmol) and N,N-diisopropylethylamine (0.53 mL, 3.02 mmol) were added, and the mixture was stirred at room temperature for 30 min. Compound 2 (0.33 mg, 1.59 mmol) was then added and stirred at room temperature for 10 min. After the reaction was completed, the reaction solution was diluted with ethyl acetate (50 mL), washed three times with saturated aqueous ammonium chloride solution (25 mL×3), dried over anhydrous sodium sulfate, and concentrated to obtain compound 20 (0.91 g, yield 87%).

[0110] 1 H NMR(300MHz,DMSO-d6)δ8.00(d,J=8.9Hz,1H,NH),7.87-7.64(m,1H,NH),7.61-7.47(m,5H,ArH),7.31-7.20(m,1H,NH),5.40(d,J=3.2Hz,1H,CH),5.00-4.88(m,3H,CH,CH2),4.65(d,J=8.3Hz,1H,CH),4.33(s,2H,CH2),4.00-3.83(m,1H,CH),3.70-3.60(m,1H,CH),3.51-3.40(m,2H,CH2),3.22-3.05(m,4H,2CH2),2.30(s,3H,CH3),2.04-1.88(m,5H,CH2,CH3),1.84-1.30(m,20H,7CH2,2CH3)。

Claims

1. A method for synthesizing a trivalent GalNAc compound, characterized in that: The synthetic route is as follows: Wherein, m is a positive integer from 1 to 10, and n is a positive integer from 1 to 10; The steps include: Step (1), using one of dichloromethane, acetonitrile or tetrahydrofuran as a reaction solvent, in the presence of a condensing agent and an organic base, the carboxyl-containing D-aminogalactose pentaacetate of formula I and the benzyloxycarbonyl-protected diamine of formula II react to obtain intermediate III; Step (2), using tetrahydrofuran or C1-C4 lower alcohol as the reaction solvent, in a hydrogen atmosphere, intermediate III is catalyzed by palladium carbon to remove the benzyloxycarbonyl protecting group to obtain intermediate IV; Step (3), using at least one of N,N-dimethylformamide, dimethyl sulfoxide or tetrahydrofuran as a reaction solvent or a mixed solvent of one selected from N,N-dimethylformamide or dimethyl sulfoxide and one selected from dichloromethane or acetonitrile, in the presence of a condensing agent and an organic base, the intermediate IV reacts with benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate to obtain a trivalent GalNAc compound with a structure as shown in Formula V.

2. The method for synthesizing a trivalent GalNAc compound according to claim 1, characterized in that: In step (1), the reaction solvent is dichloromethane or acetonitrile; in step (2), the C1-C4 lower alcohol is methanol; in step (3), the reaction solvent is tetrahydrofuran.

3. The method for synthesizing a trivalent GalNAc compound according to claim 1, characterized in that: In step (1), the molar ratio of the carboxyl-containing D-galactosamine pentaacetate to the benzyloxycarbonyl-protected diamine is 1:1 to 1:1.2, preferably 1:1.04 to 1:1.

06.

4. The method for synthesizing a trivalent GalNAc compound according to claim 1, characterized in that: In step (1), the condensing agent is selected from HATU, HBTU or HCTU; the organic base is selected from N,N-diisopropylethylamine, triethylamine, 1,8-diazabicycloundec-7-ene or N-methylmorpholine.

5. The method for synthesizing a trivalent GalNAc compound according to claim 1 or 4, characterized in that: In step (1), the molar ratio of the carboxyl-containing D-galactosamine pentaacetate to the condensing agent is 1:1.1 to 1:1.5, preferably 1:1.15 to 1:1.25; the molar ratio of the carboxyl-containing D-galactosamine pentaacetate to the organic base is 1:1.5 to 1:

3.

6. The method for synthesizing a trivalent GalNAc compound according to claim 1, characterized in that: In step (2), the molar ratio of palladium carbon to intermediate III is 0.05:1 to 0.2:1, calculated on the basis of palladium; and the loading amount of palladium in palladium carbon is 5 to 10%.

7. The method for synthesizing a trivalent GalNAc compound according to claim 1, characterized in that: In step (3), the condensing agent is selected from HATU or HBTU; the organic base is selected from N,N-diisopropylethylamine, triethylamine or N-methylmorpholine.

8. The method for synthesizing a trivalent GalNAc compound according to claim 1 or 7, characterized in that: In step (3), the molar ratio of benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate to intermediate IV is 1:3 to 1:4; the molar ratio of benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate to the condensing agent is 1:3.3 to 1:5; and the molar ratio of benzyl tris[(2-carboxyethoxy)methyl]methylcarbamate to the organic base is 1:5 to 1:

7.

9. The method for synthesizing a trivalent GalNAc compound according to claim 1, characterized in that: In step (1), the reaction time is no more than 10 minutes, preferably 5 to 10 minutes; the reaction temperature is 10 to 30°C, preferably 20 to 30°C; In step (2), the hydrogen pressure is 1 to 4 standard atmospheres; the reaction time is 4 to 6 hours; the reaction temperature is 10 to 30° C., preferably 20 to 25° C.; In step (3), the reaction time is 6 to 8 hours; the reaction temperature is 10 to 30°C, preferably 20 to 25°C.

10. The method for synthesizing a trivalent GalNAc compound according to claim 1, characterized in that: In step (1), after the reaction is completed, ethyl acetate is added to the reaction solution, and the mixture is washed with a saturated aqueous solution of ammonium chloride. The organic phase is taken, dried with anhydrous sodium sulfate, and concentrated to obtain intermediate III; In step (2), after the reaction is completed, the product is filtered and the filtrate is concentrated to obtain compound IV; In step (3), after the reaction is completed, the reaction solution is concentrated and then subjected to normal phase silica gel column chromatography using a mixed solvent of dichloromethane and methanol in a volume ratio of 20:1 to 4:1 as an eluent to purify the trivalent GalNAc compound.

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