Compatibility and Stability of a Shigella Polysaccharide—Protein Conjugate Antigen Formulated with Aluminum Salt and CpG 1018® Adjuvants

This study evaluated the formulation and stability of a quadrivalent glycoconjugate <i>Shigella</i> vaccine candidate based on four predominant strains (<i>S. flexneri</i>; <i>2a</i>, <i>3a</i>, and <i>6</i>, and <i>S. sonnei</i>) covering ~64% of global <i>Shigella</i> infections. Each glycoconjugate antigen consists of a strain-specific O-polysaccharide (O-PS) covalently linked to the carrier protein IpaB, a component of the <i>Shigella</i> type III secretion system. First, selective competitive ELISAs were developed to measure antigenicity of the four O-PS-IpaB conjugates formulated with different adjuvants (i.e., Alhydrogel^®, AH; Adju-phos^®, AP; and CpG-1018^®, CpG). Next, the monovalent <i>S. sonnei</i> O-PS-IpaB conjugate was studied to elucidate interactions with aluminum salt adjuvants (AH, AP) under different solution conditions. Third, the stability profiles of AH- or AP-adjuvanted <i>S. sonnei</i> O-PS-IpaB conjugate in various formulations (±CpG) were determined at different temperatures. Interestingly, incubation at 25 °C for 2 weeks resulted in increased antigenicity values when the antigen was bound to AP or AH, suggesting increased epitope exposure upon adjuvant binding. When bound to AP adjuvant at pH 5.8, the best glycoconjugate antigen stability was observed at elevated temperatures. The CpG adjuvant under these conditions, however, displayed incompatibility (i.e., material loss), presumably from precipitation due to lack of interaction with AP and presence of the detergent LDAO from the bulk antigen buffer. In contrast, the glycoconjugate antigen and CpG adjuvant were both bound to the AH adjuvant and stable at 2-8 °C, pH 7.0. This AH-CpG formulation of the O-PS-IpaB conjugate antigens was identified as a promising candidate for future animal immunogenicity testing.