Preparation method of linker NH-bis (tetrapolyethylene glycol-tert-butyl ester)
Through the four-step synthesis method, the reaction conditions are controlled and the use of special amine sources are solved, and the high cost and unstable problems of the existing NH-bis(tetrapolymer glycol-tert-butyl ester) preparation method is achieved, and high-efficiency and low-cost preparation is achieved, which is suitable for industrial production.
Patent Information
- Application Number
- CN202411696915.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-11-26
- Publication Date
- 2025-03-18
AI Technical Summary
The existing NH-bis(tetrapolymer glycol-tert-butyl ester) preparation methods have problems such as high cost, unstable, numerous by-products, and difficulty in purification, making it difficult to be suitable for industrial production.
The four-step method was used to synthesize NH-bis(tetrapolymer glycol-tert-butyl ester), including the synthesis of intermediates 1, 2, 3 and the preparation of the final product. By controlling the reaction conditions such as temperature and solvent, the use of a special amine source 2,4-dimethoxybenzylamine, was achieved with high yield and high purity.
It has achieved efficient, low-cost, simple and environmentally friendly preparation of NH-bis(tetrapolymer glycol-tert-butyl ester), with a total yield of 65%, a hazard-free process, suitable for industrial production, and reduced the cost of research and development of new drugs.
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Figure CN119638581A_ABST
Abstract
Description
Technical Field
[0001] The invention belongs to the field of biomedicine, and in particular relates to a method for preparing a linker NH-bis(tetraethylene glycol-tert-butyl ester). Background Art
[0002] NH-bis(tetraethylene glycol-tert-butyl ester), CAS: 2055041-41-1, contains an amino group with two terminal carboxylic acids; the amino group can easily react with carboxylic acids and activated esters to form amide bonds, and it can also react with carbonyl groups such as ketones and aldehydes under reductive amination conditions to form CN bonds; the terminal carboxylic acid can react with primary amine groups in the presence of coupling agents such as EDC, DCC, HATU, etc. to form stable amide bonds, which can be used as a linker for coupling drugs. The chemical structure is as follows:
[0003]
[0004] Radionuclide conjugated drugs (RDC) are composed of ligands, nuclides, chelators, and linkers. Radionuclide conjugated drugs are usually administered intravenously. The drugs bind to tumor cell surface antigens or proteins in the microenvironment as the blood circulates, and can exert their efficacy through internalization / phagocytosis or without endocytosis. CN115304582 discloses a group of Fap-ɑ specific tumor diagnostic imaging agents (diagnostic drugs in radionuclide conjugated drugs RDC), the composition of which is connected by a linker and a bifunctional chelate, and then coordinated and coupled with a radionuclide. This group of imaging agents is also used in diseases characterized by overexpression of fibroblast activation protein ɑ (Fap-ɑ). The patent specifically protects three coordination compounds, such as compound A, compound B and compound C, which can be quickly synthesized through the common raw material linker NH-bis (tetraethylene glycol-tert-butyl ester). Compound A is shown below:
[0005]
[0006] A search of synthesis literature, such as Org. Biomol. Chem., 2018, 16, 8579–8584, revealed the main synthesis method of NH-bis(tetraethylene glycol-tert-butyl ester), and other methods are processed or modified on this basis.
[0007]
[0008] This route uses tetraethylene glycol as raw material, expensive silver carbonate reagent, and highly explosive sodium azide synthesis process. The reduction step has a complex mechanism, and a large number of by-products make purification difficult and unstable. The total yield of the four steps is about 35%. These many defects make it unsuitable for industrial production. Summary of the invention
[0009] Purpose of the invention: The purpose of the present invention is to provide a new, efficient, low-cost, simple and environmentally friendly preparation method of the linker NH-bis(tetraethylene glycol-tert-butyl ester). The target product obtained can be used as a radionuclide drug conjugate linker, as an antibody drug conjugate linker, and as a marker for biological imaging probes to monitor the dynamic changes of molecules in vivo, etc.
[0010] Technical Solution
[0011] A method for preparing NH-bis(tetraethylene glycol-tert-butyl ester), characterized in that it is achieved by the following steps:
[0012]
[0013] Step (1), synthesis of intermediate 1:
[0014] Tert-butyl acrylate and tetraethylene glycol were dissolved in an organic solvent, potassium tert-butoxide was added at room temperature, and after the reaction was completed by TLC detection, the mixture was poured into water, extracted with petroleum ether, washed, dried, and concentrated to obtain compound 1;
[0015] The organic solvents used in the reaction are tetrahydrofuran, 2-methyltetrahydrofuran and dioxane; the molar ratio of tert-butyl acrylate to tetraethylene glycol is 1:1-1:1.5;
[0016] Step (2), Synthesis of Intermediate 2 (X=Cl):
[0017] The intermediate 2 is dissolved in dichloromethane, triethylamine is added, dichlorothionyl is added dropwise in an ice bath, and the reaction is carried out at room temperature. After the reaction is completed by TLC detection, water is added to quench the dichloromethane extraction, washing, drying, and concentrating to obtain the intermediate 2; the molar ratio of the intermediate 2 to the dichlorothionyl is 1:1-1:1.1;
[0018] Or: Synthesis of intermediate 2 (X = Br or I):
[0019] The intermediate 2 is dissolved in an organic solvent, triethylamine is added, and methanesulfonyl chloride or methanesulfonic anhydride is reacted at room temperature under an ice bath. After the reaction is completed by TLC detection, potassium bromide is added. When X=I, potassium iodide is added and the reaction is carried out at room temperature. After the reaction is completed by TLC detection, water is added to quench, tetrahydrofuran is removed by concentration, ethyl acetate is used for extraction, washing, drying, and concentration to dryness to obtain the intermediate 2; the molar ratio of the intermediate 2 to methanesulfonyl chloride or methanesulfonic anhydride is 1:1.05-1:1.1; the organic solvents for the reaction are acetonitrile and tetrahydrofuran.
[0020] Step (3) Synthesis of intermediate 3:
[0021] Dissolve intermediate 2 in an organic solvent, add intermediate 2, 2,4-dimethoxybenzylamine, potassium carbonate and a catalyst, heat to react in a nitrogen atmosphere, and after TLC detection, cool to room temperature and filter, remove the solvent under reduced pressure, extract with ethyl acetate, wash, dry, filter, and concentrate to obtain intermediate 3;
[0022] The solvents for the reaction are N,N-dimethylacetamide, N-methylpyrrolidone, and dimethyl sulfoxide; the catalysts for the reaction are sodium iodide, PEG400, PEG600, and 4-dimethylaminopyridine; the reaction temperature is 80-120° C.; the molar ratio of 2,4-dimethoxybenzylamine to intermediate 2 is 1:2.05-1:3, and the molar ratio of 2,4-dimethoxybenzylamine to potassium carbonate is 1:2-1:6; Step (4), synthesis of AN001465:
[0023] The intermediate 3 is dissolved in an organic solvent, a catalytic amount of palladium carbon is added, and the reaction is carried out in a hydrogen atmosphere at room temperature under a pressure of 15 psi. After the reaction is completed by TLC detection, the palladium carbon is recovered by filtration, the solvent is concentrated to remove, and the product is obtained by high vacuum distillation under reduced pressure; the reaction solvent is tetrahydrofuran, 2-methyltetrahydrofuran, dioxane, and isopropanol.
[0024] Beneficial effects:
[0025] 1. The synthesis of NH-bis(tetraethylene glycol-tert-butyl ester) seems simple, but the actual operation faces great difficulties. First, the intermediates 1-3 and the product (NH-bis(tetraethylene glycol-tert-butyl ester)) are all oily and difficult to crystallize, so they cannot be purified by crystallization, which has a great impact on the subsequent steps; secondly, the amino group can easily react with carboxylic acid and activated ester to form an amide bond, and it can also react with carbonyl groups such as ketones and aldehydes under reductive amination conditions to form a CN bond, that is, the active "N" leads to the generation of impurities, making it difficult to achieve a high yield.
[0026] Since each step of the present invention is an oily substance and cannot be purified, each step must have a high yield. The present invention controls the reaction conditions, such as temperature or reaction solvent, catalyst, etc., to achieve a high reaction yield. In step (3), a special amine source, namely 2,4-dimethoxybenzylamine, is used to synthesize intermediate 3 to achieve a yield of more than 79%, and the next step can be directly carried out without purification. Conventional sources such as ammonia water, benzylamine, tetramethoxybenzylamine, and dimethoxybenzylamine do not react completely, and are prone to forming monosubstituted products, quaternary ammonium salts, or incomplete reactions. The yield is low (30%-40%) and cannot be purified after post-treatment; other disubstituted methoxybenzylamines or polysubstituted methoxybenzylamines can also achieve comparable results, but the commercial price is too high and is not suitable for industrial production. In addition, 2,4-dimethoxybenzylamine is easy to dock with intermediate 2, and more importantly, it is easy to remove the protecting group;
[0027] In step (3), when intermediate 2 is used as a donor to undergo a substitution reaction with dimethoxybenzylamine to form intermediate 3, the donor must be a halide (X=Cl, Br or I); when the donor is an ester, such as methyl sulfonate (MsO), p-methylbenzenesulfonate (TsO), or 4-trifluorotoluenesulfonate (TfO), the ester can also react with dimethoxybenzylamine, but it is not feasible due to poor stability at high temperatures, resulting in reaction failure.
[0028] 2. The total yield of the four steps of the present invention is as high as 65%, there is no dangerous process, and it can be directly expanded to a ton-level scale, which is particularly suitable for industrial production, and can effectively promote the progress of new drug research and development and reduce the research and development cost of new drugs. BRIEF DESCRIPTION OF THE DRAWINGS
[0029] Figure 1 is the mass spectrometry result of AN001465;
[0030] Figure 2 Proton spectrum results of AN001465. DETAILED DESCRIPTION
[0031] Example 1 (X=Cl)
[0032] Step 1: Add tetraethylene glycol (1.14 kg, 971 mmol) and tetrahydrofuran (2.65 L) to a 5L three-necked flask, add potassium tert-butoxide (2.19 g, 19.5 mmol) in batches at 25°C, stir for 15 minutes, then drop tert-butyl acrylate (250 g, 196 mmol) and tetrahydrofuran (500 ml) into the mixed clear solution at 25°C, monitor by TLC after the addition until the reaction is complete, and then concentrate to recover tetrahydrofuran. Pour the concentrated solution into 2L of water, add 2L*3 of petroleum ether for extraction, wash with tap water (1L*2), let stand fully, and then directly concentrate to dryness to obtain a colorless oily intermediate 1 (571 g, 91%).
[0033] Step 2: Add compound 1 (300 g, 931 mmol), dichloromethane (1.5 L) and triethylamine (141 g, 1.40 mol) to a 2L three-necked flask to dissolve, then add dichlorothionyl (122 g, 1.02 mol, dichlorothionyl is added as a chlorinated compound) dropwise at 0-5°C. After the addition is complete, gradually return to room temperature for reaction. After TLC monitoring until the reaction raw materials disappear, add ice water (300 g) to quench, extract with dichloromethane (1.5 L*1), wash with tap water (500 ml*2), dry, and concentrate to obtain colorless oily intermediate 2 (295 g, 93%).
[0034] Step 3: Add 2,4-dimethoxybenzylamine (34.8g, 208mmol), chloro intermediate 2 (170g, 499mmol), potassium carbonate (96.2g, 623mmol), sodium iodide (3.12g, 20.8mmol, used as a catalyst here, the same as in the following examples), N-methylpyrrolidone (1L) to a 2L three-necked flask, heat to 110°C in a nitrogen atmosphere, react for about 10h, TLC shows that the reaction is complete. Filter, concentrate to remove the solvent, add ethyl acetate (1.2L*3) for extraction and washing, dry and filter, and concentrate the filtrate to obtain a colorless oil intermediate 3 (126g, 79%).
[0035] Step 4: Add intermediate 3 (126 g, 165 mmol), 10% palladium carbon (6.3 g, 5% W / W), isopropanol (1.37 L) to a 2L three-necked flask, replace nitrogen and hydrogen three times respectively, then react for 16 hours in a nitrogen atmosphere at a pressure of about 15 psi, and TLC shows that the reaction is complete. Filter to recover palladium carbon, concentrate to recover the solvent, and perform vacuum distillation on the concentrate to collect the fractions (110-120°C, 10 mmHg) to obtain a colorless transparent oil STD: AN001465 (96 g, 87%).
[0036] ESI-MS (m / z): 625.8;
[0037] 1 H NMR (400MHz, CDCl3) δ3.80-3.70(t,4H),3.70-3.50(m,28H),2.85-2.75(t,4H),2.60-2.40(t,4H),1.44(s,18H).
[0038] Example 2 (X=Br)
[0039] Step 1: Add tetraethylene glycol (2.28kg, 1.94mol) and tetrahydrofuran (5L) to a 10L three-necked flask, add potassium tert-butoxide (4.50g, 40.0mmol) in batches at 25°C, stir for 15 minutes, then drop tert-butyl acrylate (500g, 392mmol) and tetrahydrofuran (1.2) into the mixed clear solution at 25°C, monitor by TLC after the addition until the reaction is complete, and then concentrate to recover tetrahydrofuran. Pour the concentrated solution into 4L water, add 4L*3 of petroleum ether for extraction, wash with tap water (2L*2), let stand fully, and then directly concentrate to dryness to obtain a colorless oily intermediate 1 (1.09Kg, 87%).
[0040] Step 2: Add compound 1 (300 g, 931 mmol), tetrahydrofuran (1.5 L) and triethylamine (141 g, 1.40 mol) to a 2L three-necked flask to dissolve, then add methanesulfonyl chloride (117 g, 1.02 mol) dropwise at 0-5°C. After the addition is complete, gradually return to room temperature for reaction. After TLC monitoring until the reaction raw materials disappear, add sodium bromide (144 g, 1.40 mol) and react at 25°C for 12 h. TLC shows that the raw materials have reacted. Concentrate and recover the solvent, add ethyl acetate (1.5 L*3) for extraction, wash, dry, filter, and concentrate to obtain brominated colorless oil intermediate 2 (325 g, 91%).
[0041] Step 2: Add compound 1 (300 g, 931 mmol), acetonitrile (1.5 L) and triethylamine (141 g, 1.40 mol) to a 2L three-necked flask to dissolve, then add methanesulfonic anhydride (177 g, 1.02 mol) dropwise at 0-5°C. After the addition is complete, gradually return to room temperature for reaction. After TLC monitoring until the reaction raw materials disappear, add sodium bromide (144 g, 1.40 mol) and react at 25°C for 12 h. TLC shows that the reaction of the raw materials is complete. Concentrate and recover the solvent, add ethyl acetate (1.5 L*3) for extraction, wash, dry, filter, and concentrate to obtain brominated colorless oil intermediate 2 (308 g, 86%).
[0042] Step 3: Add 2,4-dimethoxybenzylamine (34.8g, 208mmol), bromo intermediate 2 (193, 0.50mol), potassium carbonate (96.2g, 623mmol), 4-dimethylaminopyridine (1.27g, 10.4mmol), N,N-dimethylacetamide (1L) to a 2L three-necked flask, heat to 110°C in a nitrogen atmosphere, react for about 10h, TLC shows the reaction is complete. Filter, concentrate to remove the solvent, add ethyl acetate (1.2L*3) for extraction and washing, dry and filter, and concentrate the filtrate to obtain a colorless oil intermediate 3 (118g, 73%).
[0043] Step 4: Add intermediate 3 (118 g, 152 mmol), 10% palladium carbon (5.5 g, 5% W / W), and isopropanol (1.18 L) to a 2L three-necked flask, replace nitrogen and hydrogen three times respectively, and then react for 16 hours in a nitrogen atmosphere at a pressure of about 15 psi. TLC shows that the reaction is complete. Filter to recover palladium carbon, concentrate to recover the solvent, and perform vacuum distillation on the concentrate to collect the fractions (110-120°C, 10 mmHg) to obtain a colorless transparent oil STD: AN001465 (86 g, 91%).
[0044] ESI-MS (m / z): 625.8;
[0045] 1H NMR (400MHz, CDCl3) δ3.80-3.70(t,4H),3.70-3.50(m,28H),2.85-2.75(t,4H),2.60-2.40(t,4H),1.44(s,18H).
[0046] Example 3 (X=I)
[0047] Step 1: Add tetraethylene glycol (2.28kg, 1.94mol) and tetrahydrofuran (5L) to a 10L three-necked flask, add potassium tert-butoxide (4.50g, 40.0mmol) in batches at 25°C, stir for 15 minutes, then drop tert-butyl acrylate (500g, 392mmol) and tetrahydrofuran (1.2) into the mixed clear solution at 25°C, monitor by TLC after the addition until the reaction is complete, and then concentrate to recover tetrahydrofuran. Pour the concentrated solution into 4L water, add 4L*3 of petroleum ether for extraction, wash with tap water (2L*2), let stand fully, and then directly concentrate to dryness to obtain a colorless oily intermediate 1 (1.09Kg, 87%).
[0048] Step 2: Add compound 1 (300 g, 931 mmol), tetrahydrofuran (1.5 L) and triethylamine (141 g, 1.40 mol) to a 2L three-necked flask to dissolve, then add methanesulfonyl chloride (117 g, 1.02 mol) dropwise at 0-5°C. After the addition is complete, gradually return to room temperature for reaction. After TLC monitoring until the reaction raw materials disappear, add sodium iodide (210 g, 1.40 mol) and react at 25°C for 12 h. TLC shows that the raw materials have reacted. Concentrate and recover the solvent, add ethyl acetate (1.5 L*3) for extraction, wash, dry, filter, and concentrate to obtain brominated colorless oil intermediate 2 (339 g, 95%).
[0049] Step 3: Add 2,4-dimethoxybenzylamine (34.8g, 208mmol), bromo intermediate 2 (193, 0.50mol), potassium carbonate (96.2g, 623mmol), sodium iodide (3.12g, 20.8mmol), N,N-dimethylformamide (1L) to a 2L three-necked flask, heat to 110°C in a nitrogen atmosphere, react for about 10h, TLC shows the reaction is complete. Filter, concentrate to remove the solvent, add ethyl acetate (1.2L*3) for extraction and washing, dry and filter, and concentrate the filtrate to obtain a colorless oil intermediate 3 (137g, 85%).
[0050] Step 4: Add intermediate 3 (137 g, 177 mmol), 10% palladium carbon (6.5 g, 5% W / W), and isopropanol (1.37 L) to a 2L three-necked flask, replace nitrogen and hydrogen three times respectively, and then react for 16 hours in a nitrogen atmosphere at a pressure of about 15 psi. TLC shows that the reaction is complete. Filter to recover palladium carbon, concentrate to recover the solvent, and perform vacuum distillation on the concentrate to collect the fractions (110-120°C, 10 mmHg) to obtain a colorless transparent oil STD: AN001465 (107 g, 91%).
Claims
1. A method for preparing NH-bis(tetraethylene glycol-tert-butyl ester), characterized in that Follow these steps to achieve this: The following steps are involved: Step (1), synthesis of intermediate 1: Tert-butyl acrylate and tetraethylene glycol are dissolved in an organic solvent, potassium tert-butoxide is added at room temperature, and after the reaction is completed, the mixture is poured into water, extracted with petroleum ether, washed, dried, and concentrated to obtain compound 1; Step (2), synthesis of intermediate 2, wherein X = Cl: Compound 1 was dissolved in dichloromethane, triethylamine was added, dichlorothionyl was added dropwise in an ice bath, and the reaction was carried out at room temperature. After the reaction was completed, water was added to quench the reaction, dichloromethane was extracted, washed, dried, and concentrated to obtain intermediate 2; Or the synthesis of intermediate 2, where X = Br or I: Compound 1 is dissolved in an organic solvent, triethylamine is added, and methanesulfonyl chloride or methanesulfonic anhydride is reacted at room temperature under an ice bath. After the reaction is completed, potassium bromide is added. When X=I, potassium iodide is added and the reaction is reacted at room temperature. After the reaction is completed, water is added to quench the reaction, tetrahydrofuran is removed by concentration, and the reaction is performed by extraction with ethyl acetate, washing, drying, and concentration to obtain intermediate 2. Step (3), synthesis of intermediate 3: Dissolve intermediate 2 in an organic solvent, add intermediate 2, 2,4-dimethoxybenzylamine, potassium carbonate and a catalyst, heat to react in a nitrogen atmosphere, cool to room temperature after the reaction is completed, filter, remove the solvent under reduced pressure, extract with ethyl acetate, wash, dry, filter and concentrate to obtain intermediate 3; Step (4), synthesis of AN001465: Intermediate 3 is dissolved in an organic solvent, and a catalytic amount of palladium carbon is added. The reaction is carried out in a hydrogen atmosphere at a pressure of 15 psi at room temperature. After the reaction is completed, the palladium carbon is recovered by filtration, the solvent is removed by concentration, and the product is obtained by high vacuum distillation under reduced pressure.
2. The method according to claim 1, characterized in that The organic solvent used in the reaction in step (1) is tetrahydrofuran, 2-methyltetrahydrofuran or dioxane; the molar ratio of tert-butyl acrylate to tetraethylene glycol is 1:1-1:1.
5.
3. The method according to claim 1, characterized in that In step (2), when X=Cl, the molar ratio of intermediate 2 to thionyl chloride is 1:1-1:1.1; when X=Br or I, the molar ratio of intermediate 2 to methanesulfonyl chloride or methanesulfonic anhydride is 1:1.05-1:1.1, and the organic solvent for the reaction is acetonitrile or tetrahydrofuran.
4. The method according to claim 1, characterized in that The organic solvent used in the reaction in step (3) is N,N-dimethylacetamide, N-methylpyrrolidone or dimethyl sulfoxide; the catalyst used in the reaction is sodium iodide, PEG400, PEG600 or 4-dimethylaminopyridine; the reaction temperature in step (3) is 80-120° C.; the molar ratio of 2,4-dimethoxybenzylamine to intermediate 2 is 1:2.05-1:3, and the molar ratio of 2,4-dimethoxybenzylamine to potassium carbonate is 1:2-1:
6.
5. The method according to claim 1, characterized in that The solvent for the reaction in step (4) is tetrahydrofuran, 2-methyltetrahydrofuran, dioxane or isopropanol.