Monitoring stability of meningococcal group C conjugate vaccines; correlation of physico-chemical methods and immunogenicity assays.

Two meningococcal C-CRM197 conjugates differing in oligosaccharide chain length, number of conjugation sites, conjugation chemistry and process were monitored for stability at various temperatures or after repeated freeze-thawing by physico-chemical assays. The results were compared with assessment of immunogenicity in mice, previously shown to correlate with performance of the vaccine in clinical trials. The structural stability of the oligosaccharide chains and the protein carrier varied between the two types of conjugates. Neither was adversely affected by repeated freeze-thawing but one developed conformational changes in the protein carrier, detected by optical (CD, fluorescence) and NMR spectroscopy, when incubated at 23 degrees C or above, although integrity of the oligosaccharide structure was maintained. This was not associated with any reduction in primary IgM or IgG antibody responses to meningococcal C polysaccharide. Exposure to more extreme conditions resulting in release of a substantial proportion of free saccharide from the other conjugate sample was accompanied by significant reduction in both IgG and IgM antibody responses. In conclusion, FPLC-SEC, HPAEC-PAD and NMR spectroscopy were found useful for monitoring the stability of meningococcal C-CRM197 conjugates. Although optical spectroscopy was a sensitive method for detecting modification of the protein carrier, the results did not correlate with reduced immunogenicity.