Glycoconjugation process
The present disclosure relates generally to methods of preparing glycoconjugates containing a saccharide conjugated to a carrier protein by use of stable nitroxyl radical related agent/oxidant as an oxidizing agent, to immunogenic compositions comprising such glycoconjugates, and to methods for the use of such glycoconjugates and immunogenic compositions.
1. An immunogenic composition comprising a glycoconjugate comprising a capsular polysaccharide from Streptococcus pneumoniae conjugated to a carrier protein, wherein the glycoconjugate is prepared by a process comprising:
a) reacting the capsular polysaccharide with a stable nitroxyl radical compound and an oxidant, to produce an activated capsular polysaccharide, wherein the stable nitroxyl radical compound is 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO) and the oxidant is N-chlorosuccinimide (NCS); and
b) reacting the activated capsular polysaccharide with the carrier protein comprising one or more amine groups.
2. The immunogenic composition of claim 1 , wherein the capsular polysaccharide is reacted with 0.1 to 10 molar equivalent of N-chlorosuccinimide.
3. The immunogenic composition of claim 1 , wherein the capsular polysaccharide is reacted with less than about 0.3 molar equivalent of 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO).
4. The immunogenic composition of claim 1 , wherein the capsular polysaccharide is selected from Pn-serotype 3, Pn-serotype 10A, Pn-serotype 12F, and Pn-serotype 33F capsular polysaccharides.
5. The immunogenic composition of claim 1 , wherein the carrier protein is CRM 197.
6. The immunogenic composition of claim 1 , further comprising a pharmaceutically acceptable excipient, carrier, or diluent.
7. The immunogenic composition of claim 1 , further comprising an additional antigen.
8. The immunogenic composition of claim 7 , wherein the additional antigen comprises a protein antigen or a glycoconjugate of a capsular polysaccharide derived from S. pneumonia.
9. The immunogenic composition of claim 8 , wherein the additional antigen comprises a glycoconjugate of a capsular polysaccharide selected from Pn-serotypes 1, 4, 5, 6A, 6B, 7F, 8, 9V, 11A, 14, 15B, 18C, 19A, 19F, 22F, and 23F capsular polysaccharides.
10. The immunogenic composition of claim 7 , wherein the additional antigen comprises a protein antigen or a glycoconjugate of a capsular polysaccharide derived from N. meningitidis.
11. The immunogenic composition of claim 10 , wherein the additional antigen comprises a glycoconjugate of a capsular polysaccharide selected from serotypes A, C, W135 and Y capsular polysaccharides.
12. The immunogenic composition of claim 10 , wherein the additional antigen comprises a glycoconjugate of serotypes X capsular polysaccharide.
13. The immunogenic composition of claim 7 , wherein the additional antigen comprises a glycoconjugate of a capsular polysaccharide selected from Group B Streptococcus (GBS) serotypes Ia, Ib, II, III, IV, V, VI, VII and VIII.
14. The immunogenic composition of claim 1 , further comprising an adjuvant.
15. The immunogenic composition of claim 14 , wherein the adjuvant is an aluminum-based adjuvant selected from aluminum phosphate, aluminum sulfate, and aluminum hydroxide.
16. A method of ameliorating a bacterial infection, disease or condition in a subject, comprising administering to the subject an immunologically effective amount of an immunogenic composition of claim 1 .
17. The method of claim 16 , wherein the infection, disease or condition is associated with S. pneumoniae bacteria.
18. A method of inducing a protective immune response in a subject, comprising administering to the subject an immunologically effective amount of an immunogenic composition of claim 1 .
19. An immunogenic composition comprising a glycoconjugate comprising a Pn-serotype 12F capsular polysaccharide conjugated to a carrier protein, wherein said glycoconjugate is prepared by a process comprising:
a) reacting said capsular polysaccharide with a stable nitroxyl radical compound and an oxidant, to produce an activated capsular polysaccharide, wherein said stable nitroxyl radical compound is 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO) and said oxidant is N-chlorosuccinimide; and
b) reacting the activated capsular polysaccharide with the carrier protein comprising one or more amine groups.
20. The immunogenic composition of claim 19 , further comprising an additional antigen, wherein the additional antigen comprises a glycoconjugate of a capsular polysaccharide selected from Pn-serotypes 1, 3, 4, 5, 6A, 6B, 7F, 8, 9V, 10A, 11A, 14, 15B, 18C, 19A, 19F, 22F, 23F, and 33F capsular polysaccharides.