Linaclotide solid-phase synthesis method

Abstract

The invention discloses a linaclotide solid-phase synthesis method, and belongs to the biochemical technical field. The method includes the following steps: (1) preparation of linaclotide resin; (2) cutting the linaclotide linear peptide resin obtained in the step (1), to obtain a protection group linear peptide containing Cys(Acm) and Cys(tBu); (3) oxidizing to form a first disulfide bond, to obtain a monodisulfide cyclopeptide; (4) removing an Acm protection group in the monodisulfide cyclopeptide, to obtain a dual disulfide cyclopeptide; (5) removing a tBu protection group of the dual disulfide cyclopeptide, to obtain a trisdisulfide cyclopeptide; and (6) purifying the trisdisulfide cyclopeptide by HPLC, and freeze-drying to obtain linaclotide. The process has the characteristics of simple reaction operation, easy post-processing, low cost, high yield, and considerable economic and practical values, and besides, has wide application prospect in the polypeptide drug design and synthesis field.

Description

technical field [0001] The invention belongs to the technical field of biochemistry, and in particular relates to a solid-phase synthesis method of linaclotide. Background technique [0002] Linaclotide (Linaclotide) is the first guanylate cyclase agonist (GCCA) drug so far. It was approved by the US FDA in August 2012. The trade name is Linzess. The drug is a capsule preparation and is taken orally once a day. . The compound is a polypeptide composed of 14 amino acids, including 6 cysteine ​​residues connected by 3 pairs of disulfide bonds. Molecular formula: C 59 h 79 N 15 o 21 S 6 , the molecular weight is 1526.8, and its structural sequence is as follows: [0003] [0004] H-Cys-Cys-Glu-Tyr-Cys-Cys-Asn-Pro-Ala-Cys-Thr-Gly-Cys-Tyr-OH (three pairs of disulfide bonds are 1-6, 2-10, 5-13) . [0005] Irritable bowel syndrome (Irritable Bowel Syndrome, IBS) is a common functional bowel disease, with abdominal pain or abdominal discomfort as the main symptom, which c...

AI Technical Summary

Problems solved by technology

The random oxidation of the first method will give multiple isomers, resulting in low purity of the target peptide, difficult purification, and not suitable for large-scale production

Although the second method has certain advantages over method one, it cannot completely avoid the generation of isomers

Although the third method tried a variety of protecting groups, the final target product was not obtained

These two methods belong to the method of random oxidation, and a variety of isomers will be obtained, resulting in low purity of the target peptide, difficult purification, and not suitable for large-scale production.

[0011] Chinese patent CN 104628826 A used Trt as the protecting group of Cys when synthesizing the linear peptide of linaclotide, and the formation of three pairs of disulfide bonds also adopted the method of random oxidation, but the strong oxidant iodine was selected for oxidation, although the reaction speed was accelerated, However, it is easier to produce mismatches between various disulfide bonds, resulting in the production of various isomers, which brings great difficulties to the subsequent separation and purification of target peptides

The linear peptide obtained by this method is cumbersome, costly, and low yield, and the final random oxidation is easy to cause multiple isomers, which brings difficulties to the later purification

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