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Method for preparing complexed multivalent immunogenic conjugates

A technology of immunogenicity and conjugates, which is applied in the direction of peptide preparation methods, chemical instruments and methods, carrier binding antigen/hapten components, etc., can solve the problem of reduced solubility of activated proteins and protein-PS conjugates, limited use, Adults cannot provide long-term sustained protection and other issues

Active Publication Date: 2015-08-19
UNITED STATES OF AMERICA
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Third, the resulting reduced solubility of activated proteins and protein-PS conjugates can lead to precipitation
[0031] Existing PS-based vaccines have limited use in young children and do not provide long-lasting protection in adults

Method used

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  • Method for preparing complexed multivalent immunogenic conjugates
  • Method for preparing complexed multivalent immunogenic conjugates
  • Method for preparing complexed multivalent immunogenic conjugates

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0174] Example - Materials and Methods

[0175] Materials - Tetanus toxoid (TT) was from Lederle Vaccines, Pearl River, NY and Serum Institute of India, Pune, India. Meningococcal group A and C polysaccharides (Mn A PS and Mn C PS, respectively) were from Bio-Manguinhos, Rio de Janeiro, Brazil. Mn A PS was also obtained from SynCo Bio Partners, Amsterdam, The Netherlands. Mn W135 and Y PS were obtained from Aventis Pasteur and Chiron. Hydrazine, carbohydrazide, adipic dihydrazide (ADH), acetylhydrazide, 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDC), N -(2-Hydroxyethyl)piperazine-N'-2-ethanesulfonic acid (HEPES), sodium periodate, sodium borohydride, sodium cyanoborohydride, 4-cyano-dimethylaminopyridine tetrafluoro Borate (CDAP) and 1-amino-2,3-propanediol and various amino acids were purchased from Sigma / Aldrich Chemical Company. TNBSA (2,4,6-trinitrobenzenesulfonic acid) and BCA (bicinchoninic acid) assay kits were purchased from Pierce.

[0176] M...

specific Embodiment

[0223] Subjects including reaction time, EDC concentration and concentration of amino acid and amino acid mixture Carbodiimide-catalyzed protein activation with hydrazine or hydrazide under controlled conditions

[0224] It has been reported that the reaction of proteins with hydrazine or dihydrazide catalyzed by EDC tends to lead to aggregation and precipitation of the product. Several reaction conditions were investigated for reacting 4 mg / mL TT with 0.36M hydrazine or ADH to obtain reaction products without precipitation.

[0225] Activation reactions in the absence of amino acids

[0226] Reactions were run in the presence of 12-48 mM EDC for 1-24 hours in the absence of amino acids. The reaction mixture was buffer exchanged with 30 mM NaCl, 10 mM HEPES, pH 7.5, and stored at 4°C. Some of the products of these reactions formed precipitates during the reaction or after storage of the dialyzed product at 4°C for 1-8 weeks. The degree of activation (DA; number of hydra...

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Abstract

Methods for preparing complex multivalent immunogenic conjugates that include simultaneously reacting a plurality or immunogenic-distinct polysaccharides with at least one protein to make the complex multivalent immunogenic conjugates. The simultaneous reaction involves reaction of a hydrazide group on one reactant with an aldehyde or cyanate ester group on the other reactant.

Description

field of invention [0001] The present disclosure relates to methods of making multivalent immunogenic conjugates, and conjugates made according to such methods. [0002] priority statement [0003] This application claims the benefit of US Provisional Application No. 60 / 783,490, filed March 17, 2006, which is hereby incorporated by reference in its entirety. [0004] Cross references to related applications [0005] This application is related to WO 2005 / 014037, filed August 6, 2004, and WO 2005 / 037320, filed August 6, 2004, both of which are hereby incorporated by reference in their entirety. Background of the invention [0006] Bacterial polysaccharide (PS) is a T-cell-independent antigen that induces short-term immunity in older children and adults, but usually not in younger infants. PS cannot bind to major histocompatibility complex molecules, which are required for antigen presentation to and stimulation of helper T lymphocytes. PS can stimulate B lymphocytes to pr...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/385A61K39/08A61K39/095
CPCA61K2039/62C07K1/084A61K39/116A61K2039/6037A61K39/385A61K47/6415A61K47/646A61P31/04A61P37/04Y02A50/30A61K47/50A61K39/08A61K39/095
Inventor 澈黄·罗伯特·李
Owner UNITED STATES OF AMERICA
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