Preclinical laboratory evaluation of a bivalent Staphylococcus aureus saccharide-exotoxin A protein conjugate vaccine.

A bivalent, unadjuvanted conjugate vaccine composed of Staphylococcus aureus capsular polysaccharides type 5 and 8 (T5 and T8 PS) conjugated to a novel carrier protein, the mutant nontoxic recombinant Pseudomonas aeruginosa exotoxin A (rEPA), has been the subject of recent clinical trials. A program of preclinical laboratory evaluation was carried out in support of the clinical trials conducted by the National Vaccine Evaluation Consortium. This involved physical chemical characterization and limited assessment of toxicity and immunogenicity. The carrier protein showed good stability and its conformation was essentially maintained when conjugated. The T5- and T8-rEPA conjugates were of a size range (1-3 x 10(6) g/ mol) consistent with polysaccharide conjugates. Fluorescence spectroscopy and molecular sizing showed good batch-to-batch consistency. Although all batches of final fill preparations elicited positive immune responses in the mouse model with three schedule doses of 0.25 microg of each T5/8 conjugate per dose, the mouse serum IgG response to T8 PS varied from batch to batch. Storage temperature at 37 degrees C or below or with repetitive temperature fluctuations did not significantly affect the IgG responses to T5 or T8 PS. Storage at 56 degrees C, however, diminished the mouse serum IgG response to T5 PS. The conformation of the conjugated protein and size of the conjugates correlated well with mouse immunogenicity in the thermal stability samples; significant unfolding of the protein and downshifts in molecular size of the conjugate were only observed when stored at 56 degrees C. The relatively high stability of the novel carrier protein when conjugated to large polysaccharides makes this an attractive candidate carrier protein for other conjugate vaccines. When assayed for serum IgG concentration, the bivalent T5/ 8 conjugate was found to evoke an IgG response well over the threshold value of 10 microg/ ml anti-T5 and -T8 IgG established for the ELISA immunogenicity assay.