Polysaccharide vaccine for staphylococcal infections

一种葡萄球菌、多糖的技术，应用在抗细菌药、糖衍生物、糖衍生物等方向

Inactive Publication Date: 2005-12-14
THE BRIGHAM & WOMEN S HOSPITAL INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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  • Polysaccharide vaccine for staphylococcal infections
  • Polysaccharide vaccine for staphylococcal infections
  • Polysaccharide vaccine for staphylococcal infections

Examples

Experimental program
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Effect test

Embodiment 1

[0161] Example 1: Purification of deacetylated poly-N-acetylglucosamine

[0162] It has been discovered that deacetylated poly-N-acetylglucosamine can be prepared in accordance with the present invention from any cell line expressing an intracellular attachment locus. In particular, these bacterial strains include Staphylococcus epidermidis, Staphylococcus aureus, and other Staphylococcus strains, such as Staphylococcus carves transformed with genes in the intracellular attachment locus. According to the present invention, the following specific strains can be used to purify poly-N-acetylglucosamine from inclusions: S. epidermidis RP62A (ATCC code 35984), S. epidermidis RP12 (ATCC code 35983), S. epidermidis M187, S. carnosus TM300 (pCN27), S. aureus RN4220 (pCN27) and S. aureus MN8 myxoid.

[0163] The following is a method for preparing deacetylated poly-N-acetylglucosamine from Staphylococcus species containing the intracellular attachment locus.

[0164] The starting m...

Embodiment 2

[0168] Example 2: Deacetylated poly-N-acetylglucosamine - diphtheria toxoid (DTm) Preparation of conjugate vaccines

[0169] Diphtheria toxoid is covalently bound to purified deacetylated poly-N-acetylglucosamine by reductive amination. As described in Step 1 below, aldehyde groups are first introduced onto the surface of diphtheria toxoid by treating the protein with glutaraldehyde. Subsequently, the diphtheria toxoid activated through its free amino group is reacted with deacetylated poly-N-acetylglucosamine in the presence of the reducing agent sodium cyanoborohydride, as described in step 2 below.

[0170] Step 1: Activation of diphtheria toxoid with glutaraldehyde

[0171] Using a 10kDa MWCO dialysis cartridge, 0.1M carbonate buffer (pH10) was removed at room temperature for 3 hours to dialyze 10 mg of diphtheria toxoid (4.86 mg / ml solution containing 20 mM HEPES buffer, 50 mM NaCl, PH8). When the protein solution was completely replaced by the carbonate solution, ...

Embodiment 3

[0175] Example 3: Natural poly-N-acetylglucosamine-diphtheria toxoid conjugate vaccine preparation

[0176] Natural poly-N-acetylglucosamine ( In this case, with 95-100% acetic acid substitution) covalently bound to purified diphtheria toxoid. Subsequently, CDAP-activated poly-N-acetylglucosamine was conjugated to diphtheria toxoid as described in step 2 without the need for additional spacer molecules.

[0177] Step 1: Activation of poly-N-acetylglucosamine using CDAP

[0178] 10 mg of purified poly-N-acetylglucosamine was dissolved in 150 microliters of 5M hydrochloric acid, neutralized with an equal volume of 5M sodium hydroxide, and then diluted to 1 ml with pH9.2 boric acid buffer. Prepare CDAP at a concentration of 100 mg / ml in acetonitrile and store at -20 °C for up to 1 month. Use a pipette to slowly pipette 200 μl of CDAP (containing 20 μg) into the previously mixed borate buffer containing PNAG-II (Maira et al., Infection and Immunity 2002 No. 70 pp. 4433-4440...

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Abstract

The present invention relates to compositions of staphylococcal deacetylated poly-N-acetylglucosamine (dPNAG). Deacetylated poly-N-acetylglucosamine can be isolated from natural sources or synthesized de novo. The present invention also relates to the use of deacetylated poly-N-acetylglucosamine as a vaccine to induce resistance to Staphylococcus aureus, Staphylococcus epidermidis, other related coagulase-negative or coagulase-positive staphylococci, and other carrier intracellular Active immunity to infection by organisms attached to loci. The present invention also provides a method for using the antibody targeting deacetylated poly-N-acetylglucosamine, especially a method for inducing passive immunity against similar infections.

Description

Field of Invention [0001] The present invention relates to immune-inducing polysaccharide compositions for the prevention and treatment of staphylococcal infection. The present invention also relates to methods for preparing and using polysaccharide-based antigens, related antibodies and diagnostic kits, and methods for inducing active and passive immunity by using polysaccharide materials and antibodies thereof. Background of the Invention [0002] Staphylococci are Gram-positive bacteria that normally inhabit and colonize human skin and mucous membranes. If the skin or mucous membranes are damaged during surgery or other trauma, staph bacteria have the opportunity to enter the internal tissues, causing the infection to worsen. If staphylococci proliferate locally or enter the lymphatic or blood system, serious infectious complications, such as those associated with staphylococcal bacteremia, can result. These complications include septic shock, endocarditis, arthritis, o...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/715A61K39/00A61K39/085C07K16/12
CPCA61K31/715A61K39/00A61K39/085A61K2039/6037A61K2039/627C07K16/1271G01N2400/10C08B37/0063A61P31/04A61P37/04C08B37/00A61K39/02A61K39/09A61K39/385A61K47/646A61K2039/6087
Inventor 杰拉尔德·B·皮尔汤姆斯·梅拉-利特安
Owner THE BRIGHAM & WOMEN S HOSPITAL INC
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