Electrostatic-mediated enhancement of protein antigen immunogenicity using charged TLR2-targeting lipopeptides

The low immunogenicities exhibited by most soluble proteins are in general due to the absence of any molecular signatures that are recognized by the immune system as dangerous. We show here that electrostatic binding of synthetic branched cationic or anionic lipopeptides that contain the TLR2 agonist Pam2Cys can markedly enhance protein immunogenicity. High protein-specific antibody titres in animals were achieved by vaccination with formulations containing lipopeptide and protein of opposite charge. This response was not totally dependent on electrostatic binding because vaccination with similarly charged constituents also resulted in the induction of strong, albeit lower, antibody titres. The induction of CD8+ T cell-mediated responses, however, was achieved only by vaccination with formulations containing lipopeptide and protein of opposite but not similar charge. These responses also correlated with the ability of the electrostatically associated lipopeptide to facilitate dendritic cell uptake of protein antigen and trafficking into the draining lymph node. Vaccination subsequently resulted in faster viral clearance upon pulmonary infectious challenge with a chimeric influenza virus. The improvement in protein antigen immunogenicity obtained by mixing with lipopeptide led to a 99% reduction in lung viral titres and correlates with the presence of substantial numbers of antigen-specific CD8+ T cells in lung bronchoalveolar washes.