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CN-117417401-B - Synthesis method of 2-TBS uridine monomer

CN117417401BCN 117417401 BCN117417401 BCN 117417401BCN-117417401-B

Abstract

The application relates to a 2-TBS uridine monomer synthesis method, which comprises the steps of mixing DMF, uridine and imidazole, adding DIPSCl 2 , stirring, extracting with EtOAc, washing an organic phase to obtain S1, mixing DMF, S1 and imidazole, adding TBSCl, extracting with methyl tertiary ether, washing the organic phase to obtain S2, mixing THF and S2, adding TBAF, stirring, adding EtOAc, extracting with water, washing the organic phase, drying, pulping the methyl tertiary ether to obtain S3, mixing S3 and pyridine, adding DMTrCl, stirring, adding MTBE, extracting the aqueous solution, washing the organic phase, adding citric acid aqueous solution to wash away pyridine, drying and concentrating, carrying out column chromatography to obtain S4, mixing DCM, S4, tetrazole and CTPPA, adding dichloromethane, stirring, concentrating and removing DCM, adding methyl tertiary ether, DMF and water, extracting the liquid, adding DMF and water, washing the organic phase, drying and concentrating, dropwise adding n-heptane, and crystallizing to obtain 2-TBS uridine monomer. The method has the advantages that the method effectively avoids the generation of 3' -OTBS impurities in a selective desilication mode, has simple post-treatment, can amplify production, improves yield and reduces cost.

Inventors

  • LI XIN
  • LU JIANGPING
  • CHEN FU
  • LIU SHUSEN

Assignees

  • 康羽生命科学技术(苏州)有限公司

Dates

Publication Date
20260508
Application Date
20231019

Claims (14)

  1. 1. A method for synthesizing 2-TBS uridine monomers is characterized by comprising the following synthetic routes: In Step3, tetrahydrofuran and S2 are mixed, cooled to 0-10 ℃ and stirred, TBAF is added for continuous stirring, detection is carried out after 1-5 hours, ethyl acetate is added, water extraction and delamination are carried out, an organic phase is washed for 1-3 times, drying is carried out, the solvent is removed by concentration at 20-25 ℃, and methyl tertiary butyl ether is utilized for pulping to obtain S3.
  2. 2. The method for synthesizing 2-TBS uridine monomers of claim 1, wherein DMF, uridine and imidazole are mixed in Step1, the temperature is controlled to be 20-25 ℃,1,3 dichloro-1, 3-tetraisopropyl disiloxane is added and continuously stirred, detection is carried out after 1-5 hours, after raw material consumption is complete, ethyl acetate extraction is adopted, an organic phase is washed for 1-3 times, and the solvent is concentrated, so that S1 is obtained.
  3. 3. The method for synthesizing 2-TBS uridine monomers of claim 2, wherein the molar ratio of imidazole to uridine is 3-5:1.
  4. 4. The method for synthesizing 2-TBS uridine monomers of claim 2, wherein the molar ratio of 1,3 dichloro-1, 3-tetraisopropyl disiloxane to uridine is 1.1-2:1.
  5. 5. The method for synthesizing 2-TBS uridine monomers of claim 1, wherein DMF, S1 and imidazole are stirred in Step2, temperature is controlled to be 20-25 ℃, tertiary butyl dimethyl chlorosilane is added and stirring is continued, detection is carried out after 1-5 hours, after raw material consumption is complete, methyl tertiary butyl ether is used for extraction, an organic phase is washed for 1-3 times, and S2 is obtained after solvent concentration.
  6. 6. The method for synthesizing 2-TBS uridine monomers of claim 5, wherein the molar ratio of imidazole to S1 is 3-5:1.
  7. 7. The method for synthesizing 2-TBS uridine monomers of claim 5, wherein the molar ratio of t-butyldimethyl chlorosilane to S1 is 1-2:1.
  8. 8. The method for synthesizing 2-TBS uridine monomers of claim 1, wherein the molar ratio of TBAF to S2 is 1-1.5:1.
  9. 9. The method for synthesizing 2-TBS uridine monomers of claim 1, wherein in Step4, S3 and pyridine are mixed, DMTrCl is added and stirred at a temperature of 20-25 ℃ for 1-5 hours, methyl tert-butyl ether is added, water is extracted to separate liquid, an organic phase is washed by water, pyridine is washed by adding aqueous solution of citric acid, drying and concentration are carried out, and S4 is obtained through column chromatography.
  10. 10. The method for synthesizing 2-TBS uridine monomers of claim 9, wherein the molar ratio of DMTrCl to S3 is 1-1.3:1.
  11. 11. The method for synthesizing 2-TBS uridine monomers of claim 9, wherein the volume ratio of pyridine to S3 is 5-20:1.
  12. 12. The method for synthesizing the 2-TBS uridine monomer is characterized by comprising the steps of mixing DCM, S4, tetrazole and CTPPA under the condition of no water and no oxygen in Step5, slowly adding a dichloromethane solution dissolved with S4, keeping the temperature at 20-25 ℃ and continuously stirring, detecting after 1-5 hours, concentrating and removing dichloromethane at the temperature of 10-20 ℃ until the consumption of raw materials is complete, adding methyl tertiary butyl ether, DMF and water, extracting and separating liquid, adding DMF and water, washing an organic phase for 1-3 times, drying and concentrating, and dropwise adding into n-heptane for crystallization to obtain a final product S5, namely the 2-TBS uridine monomer.
  13. 13. The method for synthesizing 2-TBS uridine monomers of claim 12, wherein the molar ratio of tetrazole to S4 is 1-2:1.
  14. 14. The method for synthesizing 2-TBS uridine monomers of claim 12, wherein the molar ratio of CTPPA to S4 is 1-2:1.

Description

Synthesis method of 2-TBS uridine monomer Technical Field The application relates to the field of pharmaceutical chemical small nucleic acid monomer raw materials, in particular to a method for synthesizing a 2-TBS uridine monomer. Background The 2-TBS uridine monomer is also called 5' -O- (4, 4-dimethoxy trityl) -2' -O- [ (tertiary butyl) dimethylsilyl ] uridine-3 ' - (2-cyanoethyl-N, N-diisopropyl) 2-TBS uridine monomer, has great potential in the research field of pharmaceutical chemistry as a small nucleic acid raw material monomer, and plays a role in the research and application field of mRNA; The current common synthesis scheme uses uridine as a starting material, firstly uses DMTrCl to protect 5'-OH, then uses TBSCl to protect 2' -OH, and finally carries out phosphating reaction to obtain a target product. However, the synthesis method has a plurality of problems, such as (1) the DMTrCl protection of 5' -OH can generate double DMTr protection impurities, crystallization is difficult to remove, column chromatography is needed, and (2) TBSCl is not selective in the protection of 2' -OH, more 3' -OTBS isomers can be obtained while the product is obtained, and the removal is difficult, so that the prior art is complex in process, the impurities are difficult to remove, and the yield is low. Disclosure of Invention The application provides a method for synthesizing 2-TBS uridine monomers, which aims to solve the problems that impurities generated in the prior art are difficult to remove, the process is complicated and the yield is low. The application provides a method for synthesizing a 2-TBS uridine monomer, which adopts the following technical scheme: A synthetic method of 2-TBS uridine monomer comprises the following synthetic route: in a specific implementation scheme, DMF, uridine and imidazole are mixed in Step1, the temperature is controlled to be 20-25 ℃,1,3 dichloro-1, 3-tetraisopropyl disiloxane is added and continuously stirred, detection is carried out after 1-5 hours, after raw material consumption is complete, ethyl acetate extraction is adopted, an organic phase is washed for 1-3 times, and the solvent is concentrated, so that S1 is obtained. In a specific embodiment, the molar ratio of imidazole to uridine is 3-5:1. In a specific embodiment, the molar ratio of 1,3 dichloro-1, 3-tetraisopropyl disiloxane to uridine is 1.1-2:1. In a specific implementation scheme, DMF, S1 and imidazole are stirred in Step2, the temperature is controlled to be 20-25 ℃, tertiary butyl dimethyl chlorosilane is added and is continuously stirred for 1-5 hours, detection is carried out, after the consumption of raw materials is complete, methyl tertiary ether is used for extraction, an organic phase is washed for 1-3 times, and after solvent is concentrated, S2 is obtained. In a specific embodiment, the molar ratio of imidazole to S1 is 3-5:1. In a specific embodiment, the molar ratio of the tert-butyldimethylchlorosilane to the S1 is 1-2:1. In a specific implementation scheme, tetrahydrofuran and S2 are mixed in Step3, cooled to 0-10 ℃, stirred and added with TBAF for continuous stirring, detection is carried out after 1-5 hours, ethyl acetate is added after raw material consumption is complete, water extraction and layering are carried out, an organic phase is washed for 1-3 times, dried, concentrated at 20-25 ℃ to remove a solvent, and methyl tertiary ether is adopted for pulping to obtain S3. In a specific embodiment, the molar ratio of TBAF to S2 is 1-1.5:1. In a specific embodiment, in Step4, S3 and pyridine are mixed, DMTrCl is added and stirred at 20-25 ℃ for 1-5 hours, methyl tert-butyl ether is added, water is used for extraction and separation, the organic phase is washed with water, aqueous solution of citric acid is added for washing away pyridine, drying and concentration are carried out, and then the S4 is obtained through column chromatography. In a specific embodiment, the molar ratio of DMTrCl to S3 is 1-1.3:1. In a specific embodiment, the volume ratio of pyridine to S3 is 5-20:1. In a specific implementation scheme, in Step5, under the anhydrous and anaerobic condition, DCM, S4, tetrazole and CTPPA are mixed and stirred, a dichloromethane solution dissolved with S4 is slowly added, the temperature is kept at 20-25 ℃ and stirring is continuously carried out, detection is carried out after 1-5 hours, the consumption of raw materials is completed, dichloromethane is concentrated and removed at 10-20 ℃ under the control of temperature, methyl tertiary ether, DMF and water are added, extraction and separation are carried out, DMF and water are added, organic phase washing is carried out for 1-3 times, drying and concentration are carried out, and the mixture is dropwise added into n-heptane for crystallization, thus obtaining the final product S5, namely the 2-TBS uridine monomer. In a specific embodiment, the molar ratio of tetrazole to S4 is 1-2:1. In a specific embodiment, the molar ratio of CT