Method for chemically synthesizing Helicobacter pylori core lipopolysaccharide oligosaccharide antigen carbohydrate chain

Patent metadata

Field	Value
Publication number	US-12018306-B2
Application number	US-202318373423-A
Country	US
Kind code	B2
Filing date	Sep 27, 2023
Priority date	May 9, 2022
Publication date	Jun 25, 2024
Grant date	Jun 25, 2024

Abstract

The disclosure discloses a method for chemically synthesizing a Helicobacter pylori core lipopolysaccharide oligosaccharide antigen carbohydrate chain, and belongs to the technical field of carbohydrate chemistry. The disclosure uses D-glucose, D-galactose and D-mannose as starting materials, which undergo a series of protection and deprotection reactions to prepare eight monosaccharide blocks. The eight monosaccharide block compounds as shown in formulas 2 to 9 undergo glycosylation reactions under catalysis of corresponding activating reagents, to prepare H. pylori lipopolysaccharide core oligosaccharide antigen fragments as shown in formula 1. The disclosure further combines the synthesized oligosaccharide fragments with a chip to make a carbohydrate chip, screens the optimal antigen fragments using patient serum, or combines the synthesized oligosaccharide fragments with carrier proteins to make glycoconjugates.

First claim

What is claimed is:

1. A method for chemically synthesizing a Helicobacter pylori lipopolysaccharide core oligosaccharide antigen, wherein an H. pylori undecasaccharide antigen shown in formula 1 is synthesized through nine monosaccharide block compounds shown in formula 2 to formula 9 and formula 24;

wherein PG 2 , PG 6 , PG 8 , PG 9 , PG 12 , PG 13 , PG 14 , PG 15 , PG 17 , PG 18 , PG 19 , PG 22 , PG 23 , PG 25 , PG 26 , PG 28 , PG 29 , PG 30 , PG 34 , PG 35 , PG 36 , and PG 37 are independently hydrogen, benzyl, 2-naphthylmethyl tert-butyldimethylsilyl, tert-butyldiphenylsilyl or triethylsilyl;
PG 1 is any of hydrogen, methyl, ethyl, tert-butyl, or benzyl;
PG 4 and PG 5 form propylidene;
PG 7 , PG 11 , PG 16 , PG 21 , PG 27 , PG 32 , PG 33 , and R 38 are selected from the group consisting of acetyl, chloroacetyl, benzoyl, pivaloyl, acetylpropionyl, 9-pentamethoxycarbonyl, 2-naphthylmethyl, and 2-p-methoxybenzyl;
PG 10 is one of acetyl, chloroacetyl, benzoyl, pivaloyl, acetylpropionyl, or 9-pentamethoxycarbonyl;
PG 20 is monochloroacetyl; PG 24 is benzyl; PG 31 is benzyl;
a Linker is —(CH 2 ) n —N—Y 1 Y 2 or —(CH 2 ) n —S—Y 1 , wherein n=1-10, and Y 1 and Y 2 are independently hydrogen, benzyl, 2-naphthylmethyl, or benzylmethoxycarbonyl; and
LG is a leaving group, and is selected from any of halogen, trichloroacetimidate, N-phenyltrifluoroacetimidate glycoside, methylthio, ethylthio, phenylthio, p-tolylthio, and dibenzyl phosphate;
the method comprising the following steps:
(a) performing a glycosylation reaction between a saccharide block donor formula 3 and a saccharide block receptor formula 2 to prepare a disaccharide compound 11 by the following synthesis route:

(b) removing the PG 7 protecting group of the disaccharide 11 selectively to prepare disaccharide 12; and performing a glycosylation reaction between the disaccharide 12 and a saccharide block donor formula 4 in the presence of an accelerator to prepare a trisaccharide compound 13 by the following synthesis route:

(c) removing the PG 11 protecting group of the trisaccharide compound 13 selectively to prepare trisaccharide 14; and performing a glycosylation reaction between the trisaccharide 14 and a saccharide block donor formula 5 in the presence of an accelerator to prepare a tetrasaccharide compound 15 by the following synthesis route:

(d) removing the PG 16 protecting group of the tetrasaccharide 15 selectively to prepare tetrasaccharide 16; and performing a glycosylation reaction between the tetrasaccharide 16 and a saccharide block donor formula 6 in the presence of an accelerator to prepare a pentasaccharide compound 17 by the following synthesis route:

(e) performing a glycosylation reaction between a saccharide block donor formula 7 and a saccharide block receptor formula 9 under the catalysis of an accelerator to prepare disaccharide 18; and removing PG 29 of the disaccharide 18 selectively to prepare disaccharide 19 and further performing a glycosylation reaction between the disaccharide 19 and a saccharide block donor formula 7 under the catalysis of an accelerator to prepare a trisaccharide module 20; and after R 38 is removed, performing a reaction of the terminal hydroxyl with trichloroacetonitrile or phenyltrifluoroacetyl chloride under an alkaline catalyst to prepare a trisaccharide donor 21 by the following synthesis route:

(f) removing the PG 21 protecting group of the pentasaccharide compound 17 selectively to prepare pentasaccharide 22; and performing a glycosylation reaction between the pentasaccharide 22 and a trisaccharide donor 21 in the presence of an accelerator to prepare an octasaccharide compound 23 by the following synthesis route:

(g) performing a glycosylation reaction between a glycosyl donor formula 7 and a receptor formula 24 under the catalysis of an accelerator to prepare disaccharide 25; and removing R 38 of the disaccharide 25 selectively, and performing a reaction of the terminal hydroxyl with trichloroacetonitrile or phenyltrifluoroacetyl chloride under an alkaline catalyst to prepare a disaccharide receptor 26 by the following synthesis route:

(h) removing the PG 20 protecting group of the octasaccharide 23 selectively to prepare octasaccharide 27; performing a glycosylation reaction between the octasaccharide 27 and a monosaccharide donor formula 8 under an accelerator to prepare a nonasaccharide compound 28; and removing the protecting group PG 32 of the nonasaccharide 28 selectively to prepare a nonasaccharide receptor 29, and performing a glycosylation reaction between the nonasaccharide receptor 29 and a glycosyl donor 26 under an accelerator to prepare undecasaccharide 30 by the following synthesis route:

and
(i) removing an acyl protecting group of the undecasaccharide 30 under an alkaline conditions and an aromatic protecting of the undecasaccharide group under a palladium on carbon/hydrogen condition to complete deprotection, resulting in a completely deprotected H. pylori lipopolysaccharide core undecasaccharide antigen as shown in formula 1.

2. The method according to claim 1 , wherein a molar ratio of the saccharide block donor formula 4 to the disaccharide 12 in step (b) is (1-2):1; and the glycosylation reaction is conducted at −10° C. to 0° C. for 3 hours to 7 hours by dissolving the glycosyl donor and the glycosyl receptor in dichloromethane, and adding molecular sieves and an activating reagent.

3. The method according to claim 1 , wherein a molar ratio of the donor formula 5 to the trisaccharide 14 in step (c) is (1-2):1; and the glycosylation reaction is conducted at −20° C. to 0° C. for 3 hours to 7 hours by dissolving the glycosyl donor and the glycosyl receptor in dichloromethane, and adding molecular sieves and an activating reagent.

4. The method according to claim 1 , wherein a molar ratio of the saccharide block donor formula 6 to the tetrasaccharide 16 in step (d) is (1-2):1; and the glycosylation reaction is conducted at −20° C. to 0° C. for 3 hours to 7 hours by dissolving the glycosyl donor and the glycosyl receptor in dichloromethane, and adding molecular sieves and an activating reagent.

5

Assignees

• Univ Jiangnan

Inventors

• Yin Jian
• Hu Jing
• Zou Xiaopeng

Classifications

• Y02P20/55

  Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups · CPC title

• Y02A50/30

  Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change · CPC title

• A61K2039/6031

  Proteins · CPC title

• A61P31/04

  Antibacterial agents · CPC title

• A61K39/385

  Haptens or antigens, bound to carriers · CPC title