Preparation and Characterization of Group A Meningococcal Capsular Polysaccharide Conjugates and Evaluation of Their Immunogenicity in Mice

Abstract

Epidemic and endemic meningitis caused by group A Neisseria meningitidis remains a problem in sub-Saharan Africa. Although group A meningococcal capsular polysaccharide (GAMP) vaccine confers immunity at all ages, the improved immunogenicity of a conjugate and its compatibility with the World Health Organization's Extended Program on Immunization offers advantages over GAMP alone. Conjugates of GAMP bound to bovine serum albumin (BSA) were synthesized, characterized, and evaluated for their immunogenicities in mice. Two methods, involving adipic acid dihydrazide (ADH) as a linker, were used. First, ADH was bound to GAMP activated with cyanogen bromide (CNBr) or with 1-cyano-4(dimethylamino)-pyridinium tetrafluoroborate (CDAP) to form GAMP_CNBrAH and GAMP_CDAPAH. These derivatives were bound to BSA by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) to form GAMP_CNBrAH-BSA and GAMP_CDAPAH-BSA. Second, ADH was bound to BSA with EDC to form AHBSA. AHBSA was bound to activated GAMP to form GAMP_CNBr-AHBSA and GAMP_CDAP-AHBSA. The yield of GAMP_CDAP-AHBSA (35 to 40%) was higher than those of the other conjugates (5 to 20%). GAMP conjugates elicited immunoglobulin G (IgG) anti-GAMP in all mice after three injections of 2.5 or 5.0 μg of GAMP: the geometric mean (GM) was highest in recipients of GAMP_CDAP-AHBSA (11.40 enzyme-linked immunosorbent assay units). Although the difference was not statistically significant, the 5.0-μg dose elicited a higher GM IgG anti-GAMP than the 2.5-μg dose. Low levels of anti-GAMP were elicited by GAMP alone. GAMP_CDAP-AHBSA elicited bactericidal activity roughly proportional to the level of IgG anti-GAMP.