Multivalent pneumococcal polysaccharide-protein conjugate composition
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.