Conjugate Vaccines from Bacterial Antigens by Squaric Acid Chemistry: A Closer Look.
By using O-SP-core (O-SPcNH2) polysaccharide, isolated from Vibrio cholera O1 lipopolysaccharide (LPS) and related synthetic substances, a detailed study of factors that affect conjugation of bacterial polysaccharides to protein carriers through squaric acid chemistry to form conjugate vaccines has been carried out. Several previously unrecognized processes that take place during the squarate labeling of the O-SPcNH2and subsequent conjugation of the formed squarate (O-SPcNH-SqOMe) have been identified. The efficiency of conjugation at pH 8.5, 9.0, and 9.5 to bovine serum albumin (BSA) and to the recombinant tetanus toxin fragment C (rTT-Hc) has been determined. The study led to a protocol for more efficient labeling of O-SPcNH2antigen with the methyl squarate group, to yield a higher-quality, more potent squarate conjugation reagent. Its use resulted in about twofold increases in conjugation efficiency (from 23-26 % on BSA to 51 % on BSA and 55 % on rTT-Hc). The spent conjugation reagent could be recovered and regenerated by treatment with MeI in the absence of additional base. The immunological properties of the experimental vaccine made from the regenerated conjugation reagent were comparable with those of the immunogen made from the parent O-SPcNH-SqOMe.
NIDDK, LBC, Section on Carbohydrates, National Institutes of Health (NIH), Bethesda, MD, 20892-0815, USA.
Peng
0000-0002-1867-0059
P
Xu
Department of Immunology and Infectious Diseases, Massachusetts General Hospital, 55 Fruit Street, Boston, MA, 02114, USA.
Meagan
M
Kelly
Laboratory of Bacterial Toxins, CBER, FDA, Silver Spring, MD, 20993, USA.
Willie F
WF
Vann
International Centre for Diarrhoeal Disease Research Bangladesh, icddr, b), Dhaka, Bangladesh.
Firdausi
F
Qadri
Department of Immunology and Infectious Diseases, Massachusetts General Hospital, 55 Fruit Street, Boston, MA, 02114, USA.|Department of Medicine, Harvard Medical School, Shattuck Street, Boston, MA, 02115, USA.|Department of Immunology and Infectious Diseases, Harvard School of Public Health, 677 Huntington Avenue, Boston, MA, 02115, USA.
Edward T
ET
Ryan
NIDDK, LBC, Section on Carbohydrates, National Institutes of Health (NIH), Bethesda, MD, 20892-0815, USA.
Pavol
0000-0001-5044-3449
P
Kováč
D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry / Q000276:immunology*; D002417:Cattle; D002771:Cholera / Q000276:immunology; D022121:Cholera Vaccines / Q000737:chemistry / Q000276:immunology*; D003503:Cyclobutanes / Q000138:chemical synthesis / Q000737:chemistry / Q000276:immunology*; D006001:Glycoconjugates / Q000138:chemical synthesis / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D051379:Mice; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000276:immunology; D013744:Tetanus Toxin / Q000737:chemistry / Q000276:immunology; D014435:Typhoid Fever / Q000276:immunology; D018074:Vaccines, Conjugate / Q000737:chemistry / Q000276:immunology; D014734:Vibrio cholerae
D000942:Antigens, Bacterial; D022121:Cholera Vaccines; D003503:Cyclobutanes; D006001:Glycoconjugates; D010446:Peptide Fragments; D013744:Tetanus Toxin; D018074:Vaccines, Conjugate; C040635:tetanus toxin fragment C; D012710:Serum Albumin, Bovine; C030042:squaric acid
O-specific polysaccharides; antigens; glycoconjugates; selective methylation; vaccines