CN101289486B - Process for synthesizing RNA monomer - Google Patents
Process for synthesizing RNA monomer Download PDFInfo
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- CN101289486B CN101289486B CN2007100395684A CN200710039568A CN101289486B CN 101289486 B CN101289486 B CN 101289486B CN 2007100395684 A CN2007100395684 A CN 2007100395684A CN 200710039568 A CN200710039568 A CN 200710039568A CN 101289486 B CN101289486 B CN 101289486B
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- China
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- monomer
- rna
- protected
- protection base
- marcwicz
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- 239000000178 monomer Substances 0.000 title claims abstract description 38
- 238000000034 method Methods 0.000 title claims abstract description 11
- 230000002194 synthesizing effect Effects 0.000 title abstract 4
- NYHBQMYGNKIUIF-UUOKFMHZSA-N Guanosine Chemical compound C1=NC=2C(=O)NC(N)=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O NYHBQMYGNKIUIF-UUOKFMHZSA-N 0.000 claims description 18
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 claims description 16
- DRTQHJPVMGBUCF-XVFCMESISA-N Uridine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-XVFCMESISA-N 0.000 claims description 14
- MIKUYHXYGGJMLM-GIMIYPNGSA-N Crotonoside Natural products C1=NC2=C(N)NC(=O)N=C2N1[C@H]1O[C@@H](CO)[C@H](O)[C@@H]1O MIKUYHXYGGJMLM-GIMIYPNGSA-N 0.000 claims description 9
- NYHBQMYGNKIUIF-UHFFFAOYSA-N D-guanosine Natural products C1=2NC(N)=NC(=O)C=2N=CN1C1OC(CO)C(O)C1O NYHBQMYGNKIUIF-UHFFFAOYSA-N 0.000 claims description 9
- 229940029575 guanosine Drugs 0.000 claims description 9
- UHDGCWIWMRVCDJ-UHFFFAOYSA-N 1-beta-D-Xylofuranosyl-NH-Cytosine Natural products O=C1N=C(N)C=CN1C1C(O)C(O)C(CO)O1 UHDGCWIWMRVCDJ-UHFFFAOYSA-N 0.000 claims description 8
- 239000002126 C01EB10 - Adenosine Substances 0.000 claims description 8
- UHDGCWIWMRVCDJ-PSQAKQOGSA-N Cytidine Natural products O=C1N=C(N)C=CN1[C@@H]1[C@@H](O)[C@@H](O)[C@H](CO)O1 UHDGCWIWMRVCDJ-PSQAKQOGSA-N 0.000 claims description 8
- 229960005305 adenosine Drugs 0.000 claims description 8
- UHDGCWIWMRVCDJ-ZAKLUEHWSA-N cytidine Chemical compound O=C1N=C(N)C=CN1[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O1 UHDGCWIWMRVCDJ-ZAKLUEHWSA-N 0.000 claims description 8
- DRTQHJPVMGBUCF-PSQAKQOGSA-N beta-L-uridine Natural products O[C@H]1[C@@H](O)[C@H](CO)O[C@@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-PSQAKQOGSA-N 0.000 claims description 7
- DRTQHJPVMGBUCF-UHFFFAOYSA-N uracil arabinoside Natural products OC1C(O)C(CO)OC1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-UHFFFAOYSA-N 0.000 claims description 7
- 229940045145 uridine Drugs 0.000 claims description 7
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 claims description 5
- 230000002378 acidificating effect Effects 0.000 claims description 5
- 239000003795 chemical substances by application Substances 0.000 claims description 5
- 229910052801 chlorine Inorganic materials 0.000 claims description 5
- 239000000460 chlorine Substances 0.000 claims description 5
- 230000026731 phosphorylation Effects 0.000 claims description 5
- 238000006366 phosphorylation reaction Methods 0.000 claims description 5
- AAAQKTZKLRYKHR-UHFFFAOYSA-N triphenylmethane Chemical compound C1=CC=CC=C1C(C=1C=CC=CC=1)C1=CC=CC=C1 AAAQKTZKLRYKHR-UHFFFAOYSA-N 0.000 claims description 5
- 150000001875 compounds Chemical class 0.000 claims description 4
- 239000000126 substance Substances 0.000 claims 1
- 108091034117 Oligonucleotide Proteins 0.000 abstract description 2
- 239000002253 acid Substances 0.000 abstract description 2
- 230000000295 complement effect Effects 0.000 abstract description 2
- 239000007791 liquid phase Substances 0.000 abstract description 2
- 238000010532 solid phase synthesis reaction Methods 0.000 abstract description 2
- 238000000746 purification Methods 0.000 abstract 5
- 230000015572 biosynthetic process Effects 0.000 abstract 1
- 125000006239 protecting group Chemical group 0.000 abstract 1
- 238000011084 recovery Methods 0.000 abstract 1
- 238000003786 synthesis reaction Methods 0.000 abstract 1
- 239000002585 base Substances 0.000 description 10
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 238000002360 preparation method Methods 0.000 description 5
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- 238000009833 condensation Methods 0.000 description 2
- 230000005494 condensation Effects 0.000 description 2
- 238000010511 deprotection reaction Methods 0.000 description 2
- PNPCRKVUWYDDST-UHFFFAOYSA-N 3-chloroaniline Chemical compound NC1=CC=CC(Cl)=C1 PNPCRKVUWYDDST-UHFFFAOYSA-N 0.000 description 1
- VGGSQFUCUMXWEO-UHFFFAOYSA-N Ethene Chemical compound C=C VGGSQFUCUMXWEO-UHFFFAOYSA-N 0.000 description 1
- 239000002841 Lewis acid Substances 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 230000006837 decompression Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- 150000007517 lewis acids Chemical class 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
Images
Classifications
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
Abstract
The invention discloses a method for synthesizing monomers of RNA. The unique 2'-protecting groups of the synthesized monomers of the RNA adopted in the method are regarded as starting points and the structure of the synthesizing monomers of the RNA is different from the structure of previous synthesizing monomers of the RNA. The method is characterized in that the protecting groups are stable in acid de-protection conditions and alkaline de-protection conditions, and after solid phase synthesis is finished, on condition that the protection group of 5'-DMT is removed and the 2'-protecting group is still on the chain, high-pressured liquid phase purification can be carried out. Regarding the products after first grade purification, the 2'-protecting groups can be completely removed in proper conditions, which can be annealed to become a double strand with another complementary strand after further purification, and the purity of the RNA oligonucleotide produced after the synthesis and purification can be above 97 percent; furthermore the purification recovery rate can be above 55 percent.
Description
The invention discloses a kind of synthetic RNA monomer methods.The RNA synthon that is adopted in this method is an origin with its unique 2 '-protection base, the RNA synthon that RNA synthon structure is different from the past.Its characteristics are that this protection base is stable under acid deprotection condition, and are also very stable under alkali property deprotection condition.After solid phase synthesis is accomplished; 2 '-protection base still can carry out the high-pressure liquid phase purifying under the situation on the chain taking off 5 '-DMT protection base; Product behind the one-level purifying can take off 2 '-protection base under suitable condition fully, can be annealed into two strands with another complementary strand after being further purified.RNA oligonucleotide purity synthetic thus and that purifying obtains can reach more than 97%, and the purifying and recovering rate can reach more than 55%.
Technical field
The present invention relates to the monomeric preparation method of a kind of RNA.
Background technology
The RNA monomer is at present by Glen Research, Transgenomic, and a few major company's monopolization such as ChemGene is produced, domestic no any RNA monomer manufacturer; The RNA monomer mass index that we rely on own technology to produce is higher than existing manufacturer production product;
Summary of the invention
The objective of the invention is to adopt unique 2 '-protection base to be origin, synthetic RNA monomer.Above-mentioned purpose of the present invention realizes through following technical proposals:
A. prepare 2 '-protect basic 1-(3-chlorobenzene)-4-piperidines alkene (CPP);
B. the uridine monomer is synthetic;
C. the cytidine monomer is synthetic;
D. the adenosine monomer is synthetic;
E. the guanosine monomer is synthetic;
Described RNA monomer compound method is characterized in that:
2 ' of employing uniqueness-protect basic 1-(3-chlorobenzene)-4-piperidines alkene (CPP) to be origin, synthetic RNA monomer.
Description of drawings
Fig. 1 is a CPP preparation method structure iron:
Fig. 2 is a uridine monomer synthetic schemes:
Fig. 3 is a cytidine monomer synthetic schemes:
Fig. 4 is an adenosine monomer synthetic schemes:
Fig. 5 is a guanosine monomer synthetic schemes:
Embodiment
The preparation method of the basic CPP of 2 '-protection:
3-chlorpromazine chloride and ethene produce 1 under the effect of lewis acid; 5-dichloro pentanone; Form N-(3-chlorobenzene)-piperidone with the condensation of 3-chloroaniline again; The latter generates dimethyl--N-(3-chlorobenzene)-piperidines ketal with the methyl alcohol condensation again, under the decompression heating condition, takes off a part methyl alcohol more at last and generates desired 1-(3-chlorobenzene)-4-piperidines alkene (CPP);
The preparation of RNA synthon:
Uridine monomer: 3 '; 5 '-hydroxy is protected with Marcwicz protection base, under acidic conditions, introduces 2 '-CPP protection base then, sloughs Marcwicz protection base subsequently; 5 '-hydroxy is with 1; 4-dimethoxy tritane chlorine (DMTrCl) is protected, and carry out phosphoramiditeization in 3 '-position with phosphorylation agent at last, makes the uridine monomer.
Cytidine monomer, adenosine monomer, the preparation of guanosine monomer: the base of cytidine/adenosine/guanosine is protected with benzoyl-earlier, and subsequent step is identical with guanosine.
Claims (2)
1. the monomeric compound method of RNA is characterized in that unique 2 '-protection base is origin, the RNA synthon that RNA synthon structure is different from the past, and compound method comprises the steps:
A. prepare 2 '-protect basic 1-(3-chlorobenzene)-4-piperidines alkene;
B. the uridine monomer is synthetic: 3 '; 5 '-hydroxy is protected with Marcwicz protection base, under acidic conditions, introduces 2 '-1-(3-chlorobenzene)-4-piperidines alkene protection base then, sloughs Marcwicz protection base subsequently; 5 '-hydroxy is with 1; 4-dimethoxy tritane chlorine is protected, and carry out phosphoramiditeization in 3 '-position with phosphorylation agent at last, makes the uridine monomer;
C. the cytidine monomer is synthetic: the base of cytidine is protected with phenmethyl earlier, and 3 ', 5 '-hydroxy is protected with Marcwicz protection base then; Under acidic conditions, introduce 2 '-1-(3-chlorobenzene)-4-piperidines alkene protection base then; Slough Marcwicz protection base subsequently, 5 '-hydroxy is with 1, and 4-dimethoxy tritane chlorine is protected; Carry out phosphoramiditeization in 3 '-position with phosphorylation agent at last, make the cytidine monomer;
D. the adenosine monomer is synthetic: the base of adenosine is protected with phenmethyl earlier, and 3 ', 5 '-hydroxy is protected with Marcwicz protection base then; Under acidic conditions, introduce 2 '-1-(3-chlorobenzene)-4-piperidines alkene protection base then; Slough Marcwicz protection base subsequently, 5 '-hydroxy is with 1, and 4-dimethoxy tritane chlorine is protected; Carry out phosphoramiditeization in 3 '-position with phosphorylation agent at last, make the adenosine monomer;
E. the guanosine monomer is synthetic: the base of guanosine is protected with phenmethyl earlier, and 3 ', 5 '-hydroxy is protected with Marcwicz protection base then; Under acidic conditions, introduce 2 '-1-(3-chlorobenzene)-4-piperidines alkene protection base then; Slough Marcwicz protection base subsequently, 5 '-hydroxy is with 1, and 4-dimethoxy tritane chlorine is protected; Carry out phosphoramiditeization in 3 '-position with phosphorylation agent at last, make the guanosine monomer.
2. the monomeric compound method of RNA as claimed in claim 1 is characterized in that, the monomeric chemical structure of the RNA of said acquisition is:
(1) uridine monomer:
(2) cytidine monomer:
(3) adenosine monomer:
(4) guanosine monomer:
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2007100395684A CN101289486B (en) | 2007-04-18 | 2007-04-18 | Process for synthesizing RNA monomer |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2007100395684A CN101289486B (en) | 2007-04-18 | 2007-04-18 | Process for synthesizing RNA monomer |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101289486A CN101289486A (en) | 2008-10-22 |
| CN101289486B true CN101289486B (en) | 2012-05-30 |
Family
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2007100395684A Active CN101289486B (en) | 2007-04-18 | 2007-04-18 | Process for synthesizing RNA monomer |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101289486B (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP2508530A1 (en) * | 2011-03-28 | 2012-10-10 | Rheinische Friedrich-Wilhelms-Universität Bonn | Purification of triphosphorylated oligonucleotides using capture tags |
| EP2712870A1 (en) * | 2012-09-27 | 2014-04-02 | Rheinische Friedrich-Wilhelms-Universität Bonn | Novel RIG-I ligands and methods for producing them |
-
2007
- 2007-04-18 CN CN2007100395684A patent/CN101289486B/en active Active
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| Publication number | Publication date |
|---|---|
| CN101289486A (en) | 2008-10-22 |
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Owner name: SHANGHAI ZHENGTI BIOTECHNOLOGY CO., LTD. Effective date: 20110824 |
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Effective date of registration: 20110824 Address after: 200086 Shanghai city Hongkou District Feihong Road No. 528 building 403 room four Applicant after: Shanghai whole biological technology Co., Ltd. Address before: 619-21 room 680, 201203 Guiping Road, Shanghai Applicant before: Shanghai Genechem Co., Ltd. |
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Owner name: SHANGHAI JIKAI GENE CHEMICAL TECHNOLOGY INC. Free format text: FORMER OWNER: SHANGHAI ZHENGTI BIOTECHNOLOGY CO., LTD. Effective date: 20120221 |
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Free format text: CORRECT: ADDRESS; FROM: 200086 HONGKOU, SHANGHAI TO: 200233 XUHUI, SHANGHAI |
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Effective date of registration: 20120221 Address after: 200233 room 619-21, Guiping Road, Shanghai, Xuhui District, 680 Applicant after: Shanghai Genechem Co., Ltd. Address before: 200086 Shanghai city Hongkou District Feihong Road No. 528 building 403 room four Applicant before: Shanghai whole biological technology Co., Ltd. |
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Address after: 200233, room 680, 619-21 Guiping Road, Shanghai, Xuhui District Patentee after: Shanghai Jikai gene Medical Technology Co.,Ltd. Address before: 200233, room 680, 619-21 Guiping Road, Shanghai, Xuhui District Patentee before: SHANGHAI GENECHEM Co.,Ltd. |