The Immunogenicity of the Tumor‐Associated Antigen Lewis^y May Be Suppressed by a Bifunctional Cross‐Linker Required for Coupling to a Carrier Protein

Abstract

A Lewis(y) (Le(y)) tetrasaccharide modified by an artificial aminopropyl spacer was synthesized by a highly convergent approach that employed a levulinoyl ester and a 9-fluorenylmethoxycarbonate for temporary protection of the hydroxy groups and a trichloroethyloxycarbonyl as an amino protecting group. The artificial aminopropyl moiety was modified by a thioacetyl group, which allowed efficient conjugation to keyhole limpet hemocyanin (KLH) modified by electrophilic 4-(maleimidomethyl)cyclohexane-1-carboxylate (MI). Mice were immunized with the KLH-MI-Le(y) antigen. A detailed analysis of sera by ELISA established that a strong immunoglobulin G (IgG) antibody response was elicited against the linker region. The use of a smaller and more flexible 3-(bromoacetamido)propionate for the attachment of Le(y) to KLH not only reduced the IgG antibody response against the linker but also led to a significantly improved immune response against the Le(y) antigen. This study shows that highly antigenic linkers suppress antibody responses to weak antigens such as self-antigens.

Identification of the Smallest Structure Capable of Evoking Opsonophagocytic Antibodies against Streptococcus pneumoniae Type 14

“…4C). Other studies have shown that anti-spacer antibodies can suppress the induction of antibodies against the carbohydrate antigen (6,22). In the present study, we found that almost all conjugates produced antibodies against the specific oligosaccharide of the conjugate that was not suppressed by the simultaneous production of anti-spacer and anti-carrier antibodies (Fig.…”

Immune recognition of tumor-associated mucin MUC1 is achieved by a fully synthetic aberrantly glycosylated MUC1 tripartite vaccine

“…Anti-MUC1 IgG, IgG1, IgG2a, IgG2b, IgG3, and IgM antibody titers were determined by ELISA as described (53). Briefly, ELISA plates (Thermo Electron) were coated with a conjugate of the MUC1 glycopeptide conjugated to BSA through a maleimide linker [BSA-MI-CTSAPDT(αGalNAc) RPAP].…”

Robust immune responses elicited by a fully synthetic three-component vaccine

“…Furthermore, the conjugation chemistry is often difficult to control resulting in conjugates with ambiguities in composition. Also, the linker moiety can elicit strong B-cell responses 5,6 . Not surprisingly, preclinical and clinical studies with carbohydrate-protein conjugates have led to results of mixed merit.…”

“…Anti-MUC1 IgG, IgG1, IgG2a, IgG2b and IgG3 antibody titers were determined by enzymelinked immunosorbent assay (ELISA), as described previously 5 . Briefly, ELISA plates (Thermo Electron Corp.) were coated with a conjugate of the MUC1 glycopeptide conjugated to BSA through a bromoacetyl linker (BSA-BrAc-MUC1).…”

“…Tumor-associated saccharides are, however, of low antigenicity, because they are self-antigens and consequently tolerated by the immune system. In addition, foreign carrier proteins such as keyhole limpet hemocyanin (KLH) and bovine serum albumin (BSA) and the linker that attach the saccharides to the carrier protein can elicit strong B-cell responses, which may lead to the suppression of antibody responses against the carbohydrate epitope 5,6 . It is clear that the successful development of carbohydrate-based cancer vaccines requires novel strategies for the more efficient presentation of tumor-associated carbohydrate epitopes to the immune system, resulting in a more efficient class switch to IgG antibodies [7][8][9][10][11][12][13][14][15][16][17] .…”