Immune stimulants (adjuvants) are essential vaccine components
however, clinically approved adjuvants are limited with the majority being derived from pathogenic components. In this study, the adjuvanting capacity of cholesterol, a natural human lipid, was explored following conjugation with peptide antigens. A structure-activity relationship study was conducted to compare linear and branched cholesterol conjugates with other lipopeptide vaccines and commercial adjuvants. Group A Streptococcus (GAS) M protein-derived J8 B-cell epitope and a universal helper T-cell epitope P25 were selected as an antigen. In addition, liposomal formulations of the cholesterol-based vaccines were also evaluated in the mouse model. Following subcutaneous and intranasal administration, conjugates comprised of cholesterol, P25 and J8 induced the highest antibody production. Linear cholesterol peptide vaccines triggered strong antibody responses that killed GAS clinical isolates as effectively as responses triggered by commercial adjuvants. The immunogenicity of the vaccines was greatly influenced by the structural arrangement of the vaccine conjugate components. The lead cholesterol conjugate was self-adjuvanting and induced the desired immune response without any exogenous immune stimulation.