Bioconjugates of
A bioconjugate of an E. coli glucosylated O4 antigen polysaccharide covalently linked to a carrier protein and compositions thereof are provided. Also provided are recombinant host cells for producing the bioconjugate, and methods of producing the bioconjugate using the recombinant host cells. The recombinant host cells contain a nucleic acid encoding a glucosyl transferase capable of modifying the E. coli O4 antigen with glucose branching to produce the glucosylated O4 antigen polysaccharide. Bioconjugates of an E. coli glucosylated O4 antigen polysaccharide described herein can be used alone or in combination with one or more additional E. coli O-antigen polysaccharides to induce antibodies against an E. coli glucosylated antigen, and to vaccinate a subject against extra-intestinal pathogenic E. coli (ExPEC).
1. A bioconjugate of an E. coli glucosylated O4 antigen polysaccharide covalently linked to a carrier protein, wherein the E. coli glucosylated O4 antigen polysaccharide comprises the structure of Formula (O4-Glc+):
wherein n is an integer of 1 to 100.
2. The bioconjugate of claim 1 , wherein the E. coli glucosylated O4 antigen polysaccharide is covalently linked to an Asn residue in a glycosylation site comprising a glycosylation consensus sequence having SEQ ID NO: 1 in the carrier protein.
3. The bioconjugate of claim 1 , wherein the carrier protein is selected from the group consisting of detoxified Exotoxin A of P. aeruginosa (EPA), E. coli flagellin (FliC), CRM197, maltose binding protein (MBP), Diphtheria toxoid, Tetanus toxoid, detoxified hemolysin A of S. aureus , clumping factor A, clumping factor B, E. coli heat labile enterotoxin, detoxified variants of E. coli heat labile enterotoxin, Cholera toxin B subunit (CTB), cholera toxin, detoxified variants of cholera toxin, E. coli Sat protein, the passenger domain of E. coli Sat protein, Streptococcus pneumoniae Pneumolysin, Keyhole limpet hemocyanin (KLH), P. aeruginosa PcrV, outer membrane protein of Neisseria meningitidis (OMPC), and protein D from non-typeable Haemophilus influenzae.
4. A composition comprising the bioconjugate of claim 1 and a pharmaceutically acceptable carrier.
5. The composition of claim 4 , comprising at least one additional antigen polysaccharide covalently linked to a carrier protein.
6. The composition of claim 5 , wherein the at least one additional antigen polysaccharide is selected from the group consisting of E. coli O1A antigen polysaccharide, E. coli O2 antigen polysaccharide, E. coli O6A antigen polysaccharide, E. coli O8 antigen polysaccharide, E. coli 015 antigen polysaccharide, E. coli 016 antigen polysaccharide, E. coli O18A antigen polysaccharide, E. coli O25B antigen polysaccharide, and E. coli O75 antigen polysaccharide.
7. The composition of claim 6 , wherein each carrier protein is EPA comprising SEQ ID NO: 3.
8. The composition of claim 6 , comprising at least the E. coli O1A, O2, glucosylated O4, O6A, O8, O15, O16, O18A, O25B, and O75 antigen polysaccharides each covalently linked to a carrier protein, wherein
(i) the E. coli O1A antigen polysaccharide comprises the structure of Formula (O1A):
(ii) the E. coli O2 antigen polysaccharide comprises the structure of Formula (O2):
(iii) the E. coli O6A antigen polysaccharide comprises the structure of Formula (O6A):
(iv) the E. coli O8 antigen polysaccharide comprises the structure of Formula (O8):
α- D -Manp3Me-(1→[3)-β- D -Manp-(1→2)-α- D -Manp-(1→2)-α- D -Manp-(1→] n ;
(v) the E. coli O15 antigen polysaccharide comprises the structure of Formula (O15):
[→2)-β- D -Galp-(1→3)-α- L -FucpNAc-(1→3)-β- D -GlcpNAc-(1→] n ;
(vi) the E. coli O16 antigen polysaccharide comprises the structure of Formula (O16):
(vii) the E. coli O18A antigen polysaccharide comprises the structure of Formula (O18A):
(viii) the E. coli O25B antigen polysaccharide comprises the structure of Formula (O25B):
and
(ix) the E. coli O75 antigen polysaccharide comprises the structure of Formula (O75):
wherein each n is independently an integer of 1 to 100.
9. The composition of claim 8 , wherein each n is an integer of 5 to 40.
10. The composition of claim 9 , wherein each n is independently an integer of 10 to 20.
11. The bioconjugate of claim 1 , wherein n is an integer of 5 to 40.
12. The bioconjugate of claim 11 , wherein n is an integer of 10 to 20.
13. The bioconjugate of claim 12 , wherein the carrier protein comprises SEQ ID NO: 3.
14. The bioconjugate of claim 11 , wherein the carrier protein comprises SEQ ID NO: 3.
15. The bioconjugate of claim 1 , wherein the carrier protein comprises SEQ ID NO: 3.
16. A composition comprising:
(i) a bioconjugate of an E. coli glucosylated O4 antigen polysaccharide covalently linked to a detoxified Exotoxin A of P. aeruginosa (EPA carrier protein) comprising SEQ ID NO: 3, wherein the E. coli glucosylated O4 antigen polysaccharide comprises the structure of Formula (O4-Glc+);
(ii) a bioconjugate of an E. coli O1A antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O1A antigen polysaccharide comprises the structure of Formula (O1A);
(iii) a bioconjugate of an E. coli O2 antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O2 antigen polysaccharide comprises the structure of Formula (O2);
(iv) a bioconjugate of an E. coli O6A antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O6A antigen polysaccharide comprises the structure of Formula (O6A);
(v) a bioconjugate of an E. coli O8 antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O8 antigen polysaccharide comprises the structure of Formula (O8);
α- D -Manp3Me-(1→[3)-β- D -Manp-(1→2)-α- D -Manp-(1→2)-α- D -Manp-(1→] n ;
(vi) a bioconjugate of an E. coli O15 antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O15 antigen polysaccharide comprises the structure of Formula (O15);
[→2)-β- D -Galp-(1→3)-α- L -FucpNAc-(1→3)-β- D -GlcpNAc-(1→] n ;
(vii) a bioconjugate of an E. coli O16 antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O16 antigen polysaccharide comprises the structure of Formula (O16);
(viii) a bioconjugate of an E. coli O25B antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O25B antigen polysaccharide comprises the structure of Formula (O25B);
(ix) a bioconjugate of an E. coli O75 antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O75 antigen polysaccharide comprises the structure of Formula (O75);
and
(x) a bioconjugate of an E. coli O18A antigen polysaccharide covalently linked to an EPA carrier protein comprising SEQ ID NO: 3, wherein the E. coli O18A antigen polysaccharide comprises the structure of Formula (O18A);
wherein each n is independently an integer of 1 to 100.
17. A composition according to claim 16 , wherein the bioconjugate of the O25B antigen polysaccharide is present in the composition at a concentration that is about 1.5-6 times-higher than the concentration of any of the other bioconjugates.
18. A composition according to claim 17 , wherein the bioconjugates of O1A:O2:glucosylated O4:O6A:O8:O15:O16:O18A:O25B:O75 are present in a ratio by weight of polysaccharide of 1:1:1:1:1:1:1:1:2:1, or 2:1:1:2:1:1:1:1:4:1.
19. A composition according to claim 17 , wherein the concentration of the bioconjugate of the 025B antigen polysaccharide is 2 to 50 μg/mL.
20. The composition of claim 16 , wherein each n is independently an integer of 5 to 40.
21. The composition of claim 20 , wherein each n is independently an integer of 10 to 20.
22. A method of inducing antibodies against an E. coli glucosylated O4 antigen in a subject, comprising administering to the subject the bioconjugate of claim 1 .
23. The method of claim 22 , wherein the subject is a human.
24. A method of vaccinating a subject against E. coli , comprising administering to the subject the bioconjugate of claim 1 .
25. The method of claim 24 , wherein the E. coli is an extra-intestinal pathogenic E. coli (ExPEC).
26. The method of claim 25 , wherein the subject is a human.
27. The method of claim 24 , wherein the subject is a human.
28. A method of inducing antibodies against E. coli in a subject, comprising administering to the subject the composition of claim 4 .
29. The method of claim 28 , wherein the subject is a human.
30. A method of vaccinating a subject against E. coli , comprising administering to the subject the composition of claim 4 .
31. The method of claim 30 , wherein the E. coli is an extra-intestinal pathogenic E. coli (ExPEC).
32. The method of claim 30 , wherein the subject is a human.
33. A method of inducing antibodies against E. coli in a subject, comprising administering to the subject the composition of claim 6 .
34. The method of claim 33 , wherein the subject is a human.
35. A method of vaccinating a subject against E. coli , comprising administering to the subject the composition of claim 6 .
36. The method of claim 35 , wherein the E. coli is an extra-intestinal pathogenic E. coli (ExPEC).
37. The method of claim 35 , wherein the subject is a human.
38. A method of inducing antibodies against E. coli in a subject, comprising administering to the subject the composition of claim 8 .
39. The method of claim 38 , wherein the subject is a human.
40. A method of vaccinating a subject against E. coli , comprising administering to the subject the composition of claim 8 .
41. The method of claim 40 , wherein the E. coli is an extra-intestinal pathogenic E. coli (ExPEC).
42. The method of claim 40 , wherein the subject is a human.