Synthesis of an S-Linked α(2→8) GD3 Antigen and Evaluation of the Immunogenicity of Its Glycoconjugate

Replacing the interglycosidic oxygen atom of oligosaccharides with a nonhydrolyzable sulfur atom has attracted significant interest because it provides opportunities for developing new glycoconjugate vaccines. Herein, a stereocontrolled and highly convergent method to synthesize a non-reducing-end inter-S-glycosidic variant of the GD3 antigen (S-linked α(2→8) GD3) that is resistant to enzymatic hydrolysis is reported. The key steps in the synthesis are a regio- and stereoselective α(2→3) sialylation of a lactoside acceptor with a C8-iodide-derivatized sialyl donor and an anomeric S-alkylation, which enable stereoselective construction of a terminal S-linked α(2→8) disialyl residue. The sulfhydryl-reactive maleimide group was used as the linker for the well-defined conjugation of these antigens to the immunogenic protein keyhole limpet hemocyanin (KLH). Groups of mice were immunized with the GD3–KLH and S-linked GD3–KLH glycoconjugates in the presence of complete Freund's adjuvant. Microarray analysis of the sera showed the promise of the S-linked GD3–KLH vaccine: it stimulated a high immunoglobulin G response against S-linked GD3 and cross-reactivity with the O-linked GD3 antigen was low. The activity of the S-linked GD3–KLH vaccine was comparable to that of the O-linked GD3–KLH vaccine, which highlighted the effectiveness of generating glycoconjugate vaccines and immunotherapies by relatively simple means.