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Synthesis method of semaglutide

A synthesis method and peptide resin technology, applied in the field of semaglutide synthesis, can solve the problems of difficult removal of peptide impurities, unfavorable purification, difficult coupling, etc., and achieve the effect of facilitating industrial scale-up and improving the purity of crude peptides

Inactive Publication Date: 2020-12-04
哈尔滨吉象隆生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, for the synthesis of semaglutide, the extension of the peptide chain is mainly achieved by coupling each amino acid unit step by step. However, many missing peptide impurities that are close to the properties of the main component will be produced during the synthesis process.
For example, alanine (Ala) or glycine (Gly) amino acid deletion with simple structure and small molecular weight produces missing peptide impurities that are difficult to remove during the purification process, which seriously affects the yield, especially the 18th and 19th peptide chains. The position amino acid is two consecutive Ala, and it is more likely to cause a single Ala deletion to produce impurities and affect the purification
[0004] In addition, the large hindrance amino acid Aib was introduced during the synthesis of semaglutide 2 After that, the N-terminal amino acid His 1 Coupling difficulties, resulting in a large amount of [des-His 1 ] impurity, if multiple re-injections are taken, it will lead to racemic impurities [D-His 1 ], which is not conducive to the subsequent purification

Method used

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  • Synthesis method of semaglutide
  • Synthesis method of semaglutide
  • Synthesis method of semaglutide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] A synthetic method of semaglutide, the steps are as follows:

[0060] 1. Resin swelling

[0061] Weigh 12.84g of Fmoc-Gly-Wang resin with a substitution degree of 0.39mmol / g and pour it into a solid-phase synthesis reactor, blow it with nitrogen, add 100mL of DMF to swell for 1 hour, and then drain the solvent.

[0062] 2. Remove Fmoc protection

[0063] Add 50 mL of piperidine:DMF (V / V) 1:3 deprotection reagent 1 to the resin, drain it after 5 minutes of reaction, add 50 mL of deprotection reagent 1 again, and drain it after 15 minutes of reaction.

[0064] 3. Washing

[0065] After deprotection, add 50mL DMF to the resin to wash the resin, drain it, add DMF again to wash, and repeat the washing for a total of 6 times. The ninhydrin test of the resin is positive.

[0066] 4. Coupling amino acids

[0067] Weigh Fmoc-Arg(pbf)-OH and 2.04g HOBT that are 3 times the mole number of Fmoc-Gly-Wang resin, add 50mL DMF to dissolve, add DIC 2.4mL under stirring, and stir for...

Embodiment 2

[0105] On the basis of Example 1, the condensing agent used when coupling Boc-His(Trt)-Aib-OH was replaced by TBTU / DIEA, and the obtained semaglutide crude peptide HPLC detection results were as follows Figure 5 As shown, the purity of semaglutide is 72.088%, and the maximum purity is 6.843%.

Embodiment 3

[0107] On the basis of Example 1, the condensing agent used when coupling Boc-His(Trt)-Aib-OH was replaced by HATU / DIEA, and the obtained semaglutide crude peptide HPLC detection results were as follows Image 6 As shown, the purity of semaglutide is 71.369%, and the maximum unmixed value is 4.609%.

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Abstract

The invention discloses a synthesis method of semaglutide. The synthesis method comprises the following steps: (1) taking Fmoc-Gly-Wang resin as a solid-phase carrier and removing an Fmoc protection group; gradually coupling amino acids from a C end to an N end in sequence; coupling to 20 amino acid R-Lys (Fmoc)-OH at the N end of a main chain; (2) removing a side chain Fmoc protection groupof the 20 amino acid at the N end of the main chain and coupling side chain amino acids in sequence; (3) removing a 20 amino acid R protection group at the N end of the main chain and coupling residual amino acids of the main chain in sequence according to a peptide sequence to obtain semaglutide full-protection peptide resin, wherein dipeptide fragments are selected as 18 -19 amino acids at the N end and 1 -2 <nd> amino acids at the N end; and (4) cracking and precipitating the semaglutide full-protection peptide resin through a cracking solution to obtain semaglutide crudepeptide. The method disclosed by the invention is simple and convenient; the prepared semaglutide has high purity and industrial production is facilitated.

Description

technical field [0001] The invention relates to the technical field of polypeptide synthesis, and more specifically relates to a method for synthesizing semaglutide. Background technique [0002] Semaglutide (Semaglutide) is a new long-acting GLP-1 analogue, its peptide sequence is as follows: His 1 -Aib 2 -Glu 3 -Gly 4 -Thr 5 -Phe 6 -Thr 7 -Ser 8 -Asp 9 -Val 10 -Ser 11 -Ser 12 -Tyr 13 -Leu 14 -Glu 15 -Gly 16 -Gln 17 -Ala 18 -Ala 19 -Lys 20 (AEEA-AEEA-γ-Glu-Octadecanedioic)-Glu 21 -Phe 22 -Ile 23 -Ala 24 -Trp 25 -Leu 26 -Val 27 -Arg 28 -Gly 29 -Arg 30 -Gly 31 -OH. From the perspective of the peptide sequence, the 2nd amino acid is unnatural α-aminoisobutyric acid (Aib), and the 20th amino acid lysine side chain of the peptide sequence has octadecanoic acid modified by polyethylene glycol (PEG) chain, so that the hydrophilicity of the entire peptide is greatly enhanced, not only can be tightly combined with albumin, resist degradation by dipepti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/605C07K1/04C07K1/06
CPCC07K14/605Y02P20/55
Inventor 冷国庆张琪谢俊侯继元杨桦于通皓杨振博王艳张凤莲景文岩黄天一孙宝伟汪莹莹关伟萍付文君
Owner 哈尔滨吉象隆生物技术有限公司
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