Multivalent pneumococcal polysaccharide-protein conjugate composition

Abstract

An immunogenic composition having 13 distinct polysaccharide-protein conjugates and optionally, an aluminum-based adjuvant, is described. Each conjugate contains a capsular polysaccharide prepared from a different serotype of Streptococcus pneumoniae (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F and 23F) conjugated to a carrier protein. The immunogenic composition, formulated as a vaccine, increases coverage against pneumococcal disease in infants and young children globally, and provides coverage for serotypes 6A and 19A that is not dependent on the limitations of serogroup cross-protection. Also described is a method for making an immunogenic conjugate comprising Streptococcus pneumoniae serotype 1 polysaccharide covalently linked to a carrier protein, the method including partial de-O-acetylation of the polysaccharide by mild hydrolysis in an alkaline pH buffer.

1 . A method for making an immunogenic conjugate comprising  Streptococcus pneumoniae  serotype 1 polysaccharide covalently linked to a carrier protein, the method comprising: 
 (a) reacting purified serotype 1 polysaccharide with an alkaline pH buffer resulting in a partially de-O-acetylated serotype 1 polysaccharide;    (b) reacting the partially de-O-acetylated serotype 1 polysaccharide with a mild acid resulting in a neutralized, partially de-O-acetylated serotype 1 polysaccharide;    (c) reacting the neutralized, partially de-O-acetylated serotype 1 polysaccharide with an oxidizing agent resulting in an activated serotype 1 polysaccharide;    (d) compounding the activated serotype 1 polysaccharide with a carrier protein;    (e) reacting the compounded, activated serotype 1 polysaccharide and carrier protein with a reducing agent resulting in a serotype 1 polysaccharide:carrier protein conjugate; and    (f) capping unreacted aldehydes in the serotype 1 polysaccharide:carrier protein conjugate resulting in an immunogenic conjugate comprising  Streptococcus pneumoniae  serotype 1 polysaccharide covalently linked to a carrier protein.    
   
   
       2 . The method of  claim 1 , further comprising purifying the activated serotype 1 polysaccharide prior to compounding with the carrier protein.  
   
   
       3 . The method of  claim 1 , further comprising co-lyophilizing the compounded, activated serotype 1 polysaccharide and carrier protein prior to reacting with the reducing agent.  
   
   
       4 . The method of  claim 1 , further comprising purifying the immunogenic conjugate.  
   
   
       5 . The method of  claim 1 , wherein the alkaline pH buffer is a bicarbonate/carbonate buffer.  
   
   
       6 . The method of  claim 1 , wherein the mild acid is acetic acid.  
   
   
       7 . The method of  claim 1 , wherein the mild acid is a low ionic strength mineral acid.  
   
   
       8 . The method of  claim 7 , wherein the low ionic strength mineral acid is hydrochloric acid.  
   
   
       9 . The method of  claim 1 , wherein the oxidizing agent is sodium periodate.  
   
   
       10 . The method of  claim 1 , wherein the carrier protein is CRM 197 .  
   
   
       11 . The method of  claim 1 , wherein the reducing agent is sodium cyanoborohydride.  
   
   
       12 . The method of  claim 1 , wherein capping unreacted aldehydes comprises reacting the serotype 1 polysaccharide:carrier protein conjugate with sodium borohydride.  
   
   
       13 . A method for making an immunogenic conjugate comprising  Streptococcus pneumoniae  serotype 1 polysaccharide covalently linked to a carrier protein, the method comprising: 
 (a) reacting purified serotype 1 polysaccharide with a bicarbonate/carbonate buffer resulting in a partially de-O-acetylated serotype 1 polysaccharide;    (b) reacting the partially de-O-acetylated serotype 1 polysaccharide with acetic acid resulting in a neutralized, partially de-O-acetylated serotype 1 polysaccharide;    (c) reacting the neutralized, partially de-O-acetylated serotype 1 polysaccharide with sodium periodate resulting in an activated serotype 1 polysaccharide;    (d) purifying the activated serotype 1 polysaccharide;    (e) compounding the activated serotype 1 polysaccharide with a carrier protein;    (f) co-lyophilizing the compounded, activated serotype 1 polysaccharide and carrier protein;    (g) reacting the co-lyophilized, activated serotype 1 polysaccharide and carrier protein with sodium cyanoborohydride resulting in a serotype 1 polysaccharide:carrier protein conjugate; and    (h) capping unreacted aldehydes in the serotype 1 polysaccharide:carrier protein conjugate with sodium borohydride resulting in an immunogenic conjugate comprising  Streptococcus pneumoniae  serotype 1 polysaccharide covalently linked to a carrier protein.    
   
   
       14 . The method of  claim 13 , further comprising purifying the immunogenic conjugate.  
   
   
       15 . The method of  claim 13 , wherein the carrier protein is CRM 197 .  
   
   
       16 . A method for making an activated  Streptococcus pneumoniae  serotype 1 polysaccharide, the method comprising: 
 (a) reacting purified serotype 1 polysaccharide with an alkaline pH buffer resulting in a partially de-O-acetylated serotype 1 polysaccharide;    (b) reacting the partially de-O-acetylated serotype 1 polysaccharide with a mild acid resulting in a neutralized, partially de-O-acetylated serotype 1 polysaccharide; and    (c) reacting the neutralized, partially de-O-acetylated serotype 1 polysaccharide with an oxidizing agent resulting in an activated serotype 1 polysaccharide.    
   
   
       17 . The method of  claim 16 , further comprising purifying the activated serotype 1 polysaccharide following oxidation.  
   
   
       18 . The method of  claim 16 , wherein the alkaline pH buffer is a bicarbonate/carbonate buffer.  
   
   
       19 . The method of  claim 16 , wherein the mild acid is acetic acid.  
   
   
       20 . The method of  claim 16 , wherein the oxidizing agent is sodium periodate.