A method for the synthesis of a polyethylene glycol drug modifier

Abstract

The application discloses a synthesis method of a polyethylene glycol drug modifier, which comprises the following steps: S1: polyethylene glycol is mixed with dry organic solvent in a reactor, and then tert-butyl acrylate is added dropwise after adding an alkali and keeping for a certain time; tBuOOCCH2CH2-PEGn-CH2CH2COOtBu with a purity greater than 99% is obtained; S2: tBuOOCCH2CH2-PEGn-CH2CH2COOtBu obtained in S1 is dissolved with an organic solvent, and an alkali aqueous solution is added to carry out a homogeneous saponification reaction; HOOCCH2CH2OPEGn-CH2CH2COOtBu is obtained by an acid-alkali treatment; S3: HOOCCH2CH2OPEGn-CH2CH2COOtBu obtained in S2 is dissolved with anhydrous organic solvent, and an organic alkali and diphenyl phosphinic azide are added to react at room temperature, and then a reflux reaction is carried out by heating.

Description

Technical Field

[0001] This invention relates to the field of drug modifier preparation technology, and specifically to a method for synthesizing a polyethylene glycol drug modifier. Background Technology

[0002] Polyethylene glycol (PEG) and its derivatives possess the following excellent properties: ① They are water-soluble and soluble in most organic solvents. ② PEG products are non-toxic, have low immunogenicity, and their biocompatibility has been certified by the US FDA. ③ PEG has many different molecular weights and a wide range of applications. ④ PEG imparts many excellent properties to its modified small biological molecules.

[0003] With the development of drug research, the shortcomings of many drugs have gradually become apparent. For example, although some drugs have good efficacy, high specificity, and low toxicity, they also have problems such as short half-life, poor stability, poor water solubility, immunogenicity, and rapid renal clearance. Polyethylene glycol-modified protein drugs or small molecule drugs create a spatial barrier around their molecules, maintaining protein activity, reducing immunogenicity, prolonging the in vivo half-life, increasing the drug molecular weight, and reducing renal clearance.

[0004] With the development of biotechnology, protein-based biopharmaceuticals are being developed and applied more and more widely due to their advantages such as high activity, strong specificity, well-defined biological functions, and low toxicity. Modifiers for polyethylene glycol (PEG) are constantly being improved and innovated, and specific linkages between different drugs are being explored. The development of PEG derivatives has progressed through the first generation of random modification, the second generation of specific and functional modification, and the third generation of branched structure applications. Their applications have also expanded from simple drug modification to areas such as biosensing and drug delivery.

[0005] The application of polyethylene glycol (PEG) in modern pharmaceutical manufacturing began in the 1970s. Currently, nearly 20 PEG-modified drugs are on the market globally, with a market size exceeding $10 billion. It remains a hot industry, and optimizing synthetic processes is a crucial step to reduce costs, improve the purity of modifiers, and protect the environment.

[0006] Based on the above, this invention proposes a method for synthesizing polyethylene glycol drug modifiers, which can effectively solve the above problems. Summary of the Invention

[0007] The purpose of this invention is to provide a method for synthesizing polyethylene glycol (PEG) drug modifiers. The method employs a novel synthetic route, using diphenyl azidophosphate to synthesize amino groups, and the amino groups can be protected during the synthesis process, thereby shortening the synthetic route. The short synthetic route is simple to operate, reduces the synthesis cycle, and thus significantly lowers the synthesis cost.

[0008] To solve the above technical problems, the technical solution provided by the present invention is as follows:

[0009] A method for synthesizing a polyethylene glycol drug modifier includes the following steps:

[0010] S1: Polyethylene glycol and dry organic solvent are mixed in a reactor, alkali is added and the mixture is kept at a certain temperature for a certain period of time, and then tert-butyl acrylate is added dropwise to the above system to carry out the reaction; after the reaction is completed, the solvent and excess tert-butyl acrylate are removed by vacuum distillation; the crude product is obtained by vacuum distillation or column chromatography to obtain tBuOOCCH2CH2-PEGn-CH2CH2COOtBu with a purity greater than 99%;

[0011] S2: Dissolve tBuOOCCH2CH2-PEGn-CH2CH2COOtBu obtained in S1 in an organic solvent, add an alkaline aqueous solution and carry out a homogeneous saponification reaction under heat preservation; determine the product content by high performance liquid chromatography to confirm that the raw material reaction is complete; when there is no raw material, stop the reaction and obtain HOOCCH2CH2OPEGn-CH2CH2COOtBu by acid-base method.

[0012] S3: Dissolve HOOCCH2CH2OPEGn-CH2CH2COOtBu obtained in S2 in an anhydrous organic solvent, add an organic base and diphenyl azidophosphate, react at room temperature, and then heat to reflux; add tert-butanol, benzyl alcohol or water to the above reaction system to continue the reaction, and purify to obtain R2-CH2CH2OPEGn-CH2CH2COOtBu;

[0013] S4: Remove the protecting group of the carboxyl group from R2-CH2CH2OPEGn-CH2CH2COOtBu obtained in S3 by acid or base treatment, recrystallize, filter, and remove the solvent to obtain R2-CH2CH2OPEGn-CH2CH2COOH with an additional ethylene glycol.

[0014] The synthesis method of the polyethylene glycol drug modifier of the present invention employs a novel synthetic route, using diphenyl azidophosphate to synthesize the amino group, and the amino group can be protected during the synthesis process, thereby shortening the synthetic route. The synthetic route is short, the operation is simple, and the synthesis cycle is reduced, thus significantly lowering the synthesis cost.

[0015] Preferably, in step S1, the step of adding alkali and keeping the temperature warm for a certain period of time is 25-35 minutes after adding alkali and keeping the temperature warm.

[0016] Preferably, in step S1, the organic solvent is any one or a mixture of two or more of dichloromethane, tetrahydrofuran, sodium alkali, sodium tert-butoxide, and potassium tert-butoxide.

[0017] Preferably, in step S2, the alkali in the alkaline aqueous solution is any one or a mixture of two or more of lithium hydroxide, sodium hydroxide, and potassium hydroxide.

[0018] Preferably, in step S2, the organic solvent is any one or a mixture of two or more of methanol, acetonitrile, and 1,4-dioxane.

[0019] Preferably, in step S3, R2 in R2-CH2CH2OPEGn-CH2CH2COOtBu is tert-butyloxycarbonyl, benzyloxycarbonyl, or hydrogen.

[0020] Preferably, in step S3, when R2 is hydrogen, amino protection is performed under alkaline conditions, and Fmoc-OSu is selected as the feed reaction to obtain a compound in which R2 is fluorene methyloxycarbonyl.

[0021] Preferably, in step S3, the organic solvent is any one or a mixture of two or more of toluene, tetrahydrofuran, and 1,4-dioxane.

[0022] Preferably, in step S4, the acid is hydrochloric acid or trifluoroacetic acid.

[0023] Preferably, in step S4, the alkali is any one or a mixture of two or more of lithium hydroxide, sodium hydroxide, and potassium hydroxide.

[0024] The present invention also provides a polyethylene glycol drug modifier prepared by the synthesis method of polyethylene glycol drug modifier as described above.

[0025] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0026] In recent years, there has been a lot of research on this type of linker, and there are two relatively similar synthetic routes: Route 1 and Route 2.

[0027] Route 1:

[0028]

[0029] In the synthesis process of Route 1, both methanesulfonyl chloride and sodium azide are highly toxic reagents. In addition, sodium azide is an explosive and highly dangerous substance, which limits its use in industrial production. Furthermore, the synthesis route is relatively long.

[0030] Route 2:

[0031]

[0032] Diethylene glycolamine is used as a starting material, and the next step of synthesis is carried out through amino protection. In actual synthesis, impurity amines are generated, making product purification difficult. Furthermore, the synthetic route is long and costly.

[0033] Route 3:

[0034]

[0035] The present invention, Route 3, mainly involves the synthesis of compounds of the Boc / Cbz / Fmoc-PEGn-CH2CH2COOH class, where n is the number of ethylene glycols.

[0036] The synthesis method of the polyethylene glycol drug modifier of this invention employs a novel synthetic route, using diphenyl azidophosphate to synthesize the amino group, and the amino group can be protected during the synthesis process, thus shortening the synthetic route. The short synthetic route is simple to operate, reducing the synthesis cycle and significantly lowering the synthesis cost. It avoids the highly toxic and explosive sodium azide in route one, improving both production safety and yield. Compared to route two, the shortened synthetic route avoids the problems of amino group contamination and difficulty in purification during chain extension, making this synthesis simple and highly suitable for industrial production. Furthermore, when synthesizing the same number of ethylene glycol linkers, this invention uses one less ethylene glycol unit as raw material, thus saving costs in terms of raw materials. Attached Figure Description

[0037] Figure 1 The 1H NMR spectrum of the modifier Fmoc-NHPEG3-CH2CH2COOH;

[0038] Figure 2 The image shows the 1H NMR spectrum of the modifier Cbz-NHPEG12-CH2CH2COOH. Detailed Implementation

[0039] To enable those skilled in the art to better understand the technical solutions of the present invention, the preferred embodiments of the present invention are described below in conjunction with specific examples, but these should not be construed as limiting the present patent.

[0040] Unless otherwise specified, the test methods or experimental methods described in the following examples are conventional methods; unless otherwise specified, the reagents and materials are obtained from conventional commercial sources or prepared by conventional methods.

[0041] Example 1:

[0042] S1: Weigh 106 g (1 mol) of diethylene glycol into a 1 L reactor equipped with a mechanical stirrer, add 300 mL of anhydrous tetrahydrofuran, and add potassium tert-butoxide (2.24 g, 0.02 mol) while stirring at 25 °C. Stir at this temperature for 0.5 h. Then slowly add tert-butyl acrylate (268.8 g, 2.1 mol), and react for 6 hours. The endpoint of the reaction is determined by TLC. Ethyl acetate is added to the reaction system and washed twice with water. The ethyl acetate phase is dried, filtered, and evaporated to dryness to obtain a crude product. The crude product is then subjected to vacuum distillation using an oil pump to obtain 315 g of tBuOOCCH2CH2-PEG2-CH2CH2COOtBu. The purity was determined by gas chromatography to be 99.27%, and the yield was 87%.

[0043] S2: Take 181 g (0.5 mol) of tBuOOCCH2CH2-PEG2-CH2CH2COOtBu and place it in a 2 L three-necked flask. Add 900 mL of methanol and maintain the temperature at 15°C. Slowly add a solution of sodium hydroxide (24 g, 0.6 mol) dissolved in 40 mL of water. The system reacts homogeneously for 3 hours. The reaction is confirmed to be complete by TLC or the endpoint is determined by high performance liquid chromatography. The solution is removed under vacuum using a water pump, and the recovered methanol is reused for the next batch of reaction. The crude product is dissolved in water under alkaline conditions and washed three times with ethyl acetate to remove impurities. The aqueous phase is adjusted to pH 4 with hydrochloric acid, and the product is extracted three times with dichloromethane. The dichloromethane phase is collected, washed once with water, dried, and concentrated to obtain 93.3 g of HOOCCH2CH2-PEG2-CH2CH2COOtBu (yield 61%).

[0044] S3: Under nitrogen protection, weigh 76.5 g (0.25 mol) of HOOCCH2CH2-PEG2-CH2CH2COOtBu into a 2000 mL three-necked flask, add 50.5 g (0.5 mol) of triethylamine and 165 g (0.6 mol) of diphenyl azidophosphate into 800 mL of tetrahydrofuran, and react for 3 hours. Add 300 mL of water to the reaction system, heat to reflux, and continue the reaction for 2 hours. Remove the tetrahydrofuran by rotary evaporation, adjust the pH of the aqueous phase to 7 with hydrochloric acid, and extract impurities 6 times with dichloromethane. Adjust the pH of the aqueous phase to alkaline with sodium bicarbonate. Add the aqueous phase into a 2000 mL three-necked flask, add 600 mL of tetrahydrofuran, and add Fmoc-OSu (92.8 g, 0.275 mol) in portions to the above system, and react overnight at room temperature. The next day, the reaction solution was extracted twice with ethyl acetate, the ethyl acetate phase was collected, washed three times with water, and the organic phase was dried, filtered, and evaporated to dryness to obtain 99.8 g of Fmoc-NHCH2CH2-PEG2-CH2CH2COOtBu, also known as Fmoc-NHPEG3-CH2CH2COOtBu (yield 80%).

[0045] S4: Weigh 99.8 g (0.2 mol) of Fmoc-NHCH2CH2-PEG2-CH2CH2COOtBu into a 1000 mL single-necked flask, add 100 mL of dichloromethane and 100 mL of trifluoroacetic acid. React the solution at room temperature for 6 hours, and confirm the reaction is complete by TLC. The reaction solution is then desolvated under vacuum using a water pump, and residual trifluoroacetic acid is removed using an oil pump. The crude product is dissolved in 1500 mL of ethyl acetate and 500 mL of petroleum ether, recrystallized, and filtered to obtain 81.5 g of a white solid, Fmoc-NHCH2CH2-PEG2-CH2CH2COOH, with a purity of 99.3% (yield 92%).

[0046] Example 2:

[0047] S1: Weigh 100.4 g (0.2 mol) of undecylethylene alcohol into a 1 L reactor equipped with a mechanical stirrer, add 300 mL of anhydrous tetrahydrofuran, and add potassium tert-butoxide (0.448 g (0.004 mol) while stirring at 25 °C. Stir at this temperature for 0.5 h. Then slowly add tert-butyl acrylate (53.76 g (0.42 mol) dropwise, and react for 6 h. The endpoint of the reaction is determined by TLC. Ethyl acetate is added to the reaction system and washed twice with water. The ethyl acetate phase is dried, filtered, and evaporated to dryness to obtain the crude product. Purification by column chromatography yields 128.9 g of tBuOOCCH2CH2-PEG11-CH2CH2COOtBu, with a purity of 99.51% and a yield of 85% as determined by ELSD.

[0048] S2: Take 75.8 g (0.1 mol) of tBuOOCCH2CH2-PEG11-CH2CH2COOtBu and place it in a 2 L three-necked flask. Add 400 mL of methanol and maintain the temperature at 15°C. Slowly add a solution of sodium hydroxide (4.8 g, 0.12 mol) dissolved in 10 mL of water. The system reacts homogeneously for 3 hours. The reaction is confirmed to be complete by TLC or the endpoint is determined by high performance liquid chromatography. The solution is removed under vacuum using a water pump, and the methanol is recovered for reuse in the next batch of reaction. The crude product is dissolved in water under alkaline conditions and washed three times with ethyl acetate to remove impurities. The aqueous phase is adjusted to pH 4 with hydrochloric acid, and the product is extracted three times with dichloromethane. The dichloromethane phase is collected, washed five times with water, dried, and concentrated to obtain 44.2 g of HOOCCH2CH2-PEG11-CH2CH2COOtBu (yield 63%).

[0049] S3: Under nitrogen protection, 35.1 g (0.05 mol) of HOOCCH2CH2-PEG11-CH2CH2COOtBu was weighed and placed in a 1000 mL three-necked flask. Triethylamine (10.1 g, 0.1 mol) and diphenyl azidophosphate (16.5 g, 0.06 mol) were added to 350 mL of toluene and refluxed for 3 hours. The reaction system was cooled to room temperature, and benzyl alcohol was added to the system, and the reaction was continued for 2 hours. The solvent in the reaction solution was removed by rotary evaporation, and the crude product was dissolved in ethyl acetate and washed three times with water. The ethyl acetate phase was collected, dried, filtered, and evaporated to dryness to obtain 34.3 g of Cbz-NHCH2CH2-PEG11-CH2CH2COOtBu, also known as Cbz-NHPEG12-CH2CH2COOtBu (yield 85%).

[0050] S4: Weigh 24.2 g (0.03 mol) of Cbz-NHCH2CH2-PEG11-CH2CH2COOtBu into a 500 mL single-necked flask, add 25 mL of dichloromethane and 25 mL of trifluoroacetic acid. React the reaction solution at room temperature for 6 hours, and confirm the completion of the reaction by TLC. The reaction solution is then desolvated under vacuum using a water pump, and the residual trifluoroacetic acid is removed using an oil pump. The crude product is dissolved in 1500 mL of ethyl acetate and 500 mL of petroleum ether, recrystallized in a -40°C freezer, filtered to obtain a white solid, dried under vacuum, and desolvated at 5°C to obtain 21 g of Cbz-NHCH2CH2-PEG11-CH2CH2COOH (also known as Cbz-NHPEG12-CH2CH2COOH), with a purity of 99.1% (yield 93.2%).

[0051] The above are merely preferred embodiments of the present invention. It should be noted that the above preferred embodiments should not be considered as limitations on the present invention, and the scope of protection of the present invention should be determined by the scope defined in the claims. For those skilled in the art, several improvements and modifications can be made without departing from the spirit and scope of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for synthesizing a polyethylene glycol drug modifier, characterized in that, Includes the following steps: S1: Polyethylene glycol and dry organic solvent are mixed in a reactor, alkali is added and the mixture is kept at a certain temperature for a certain period of time, and then tert-butyl acrylate is added dropwise to the above system to carry out the reaction; after the reaction is completed, the solvent and excess tert-butyl acrylate are removed by vacuum distillation; the crude product is obtained by vacuum distillation or column chromatography to obtain tBuOOCCH2CH2-PEGn-CH2CH2COOtBu with a purity greater than 99%; S2: Dissolve tBuOOCCH2CH2-PEGn-CH2CH2COOtBu obtained in S1 in an organic solvent, add an alkaline aqueous solution and carry out a homogeneous saponification reaction under heat preservation; determine the product content by high performance liquid chromatography to confirm that the raw material reaction is complete; when there is no raw material, stop the reaction and obtain HOOCCH2CH2OPEGn-CH2CH2COOtBu by acid-base method. S3: Dissolve HOOCCH2CH2OPEGn-CH2CH2COOtBu obtained in S2 in an anhydrous organic solvent, add an organic base and diphenyl azidophosphate, react at room temperature, and then heat to reflux; add tert-butanol, benzyl alcohol or water to the above reaction system to continue the reaction, and purify to obtain R2-NH-CH2CH2OPEGn-CH2CH2COOtBu; S4: The carboxyl protecting agent of R2-NH-CH2CH2OPEGn-CH2CH2COOtBu obtained in S3 is removed by acid or base treatment, recrystallization, filtration, and solvent removal to obtain R2-NH-CH2CH2OPEGn-CH2CH2COOH with an additional ethylene glycol. In the R2-NH-CH2CH2OPEGn-CH2CH2COOH group, R2 is a tert-butyloxycarbonyl, benzyloxycarbonyl, or fluorenemethyloxycarbonyl substituent. When R2 is a substituent of tert-butyloxycarbonyl and benzyloxycarbonyl, i.e. Boc and Cbz, the synthesis process is as follows: When R2 is a substituent of fluorenemethyloxycarbonyl, i.e., Fmoc, the synthesis process is as follows: ； Where n takes the value of an integer from 2 to 11.

2. The method for synthesizing the polyethylene glycol drug modifier according to claim 1, characterized in that, In step S1, the step of adding alkali and keeping the temperature warm for a certain period of time is referred to as adding alkali and keeping the temperature warm for 25-35 minutes.

3. The method for synthesizing the polyethylene glycol drug modifier according to claim 1, characterized in that, In step S2, the alkali in the alkaline aqueous solution is sodium hydroxide.

4. The method for synthesizing the polyethylene glycol drug modifier according to claim 1, characterized in that, In step S1, the organic solvent is tetrahydrofuran, and the base is potassium tert-butoxide.

5. The method for synthesizing the polyethylene glycol drug modifier according to claim 1, characterized in that, In step S3, the organic solvent is toluene.

6. The method for synthesizing the polyethylene glycol drug modifier according to claim 1, characterized in that, In step S4, the acid is hydrochloric acid or trifluoroacetic acid.

7. The method for synthesizing the polyethylene glycol drug modifier according to claim 1, characterized in that, In step S4, the alkali is any one or a mixture of two or more of lithium hydroxide, sodium hydroxide, and potassium hydroxide.

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