The LTR72 mutant of heat-labile enterotoxin of Escherichia coli enhances the ability of peptide antigens to elicit CD4+ T cells and secrete gamma interferon after coapplication onto bare skin

Application of antigens with an adjuvant onto bare skin is a needle-free and pain-free immunization procedure that delivers antigens to the immunocompetent cells of the epidermis. We tested here the immunogenicity and adjuvanticity of two mutants of heat-labile enterotoxin (LT) of Escherichia coli, LTK63 and LTR72. Both mutants were shown to be immunogenic, inducing serum and mucosal antibody responses. The application of LTK63 and LTR72 to bare skin induced significant protection against intraperitoneal challenge with a lethal dose of LT. In addition, both LT mutants enhanced the capacity of peptides TT:830-843 and HA:307-319 (representing T-helper epitopes from tetanus toxin and influenza virus hemagglutinin, respectively) to elicit antigen-specific CD4(+) T cells after coapplication onto bare skin. However, only mutant LTR72 was capable of stimulating the secretion of high levels of gamma interferon. These findings demonstrate that successful skin immunization protocols require the selection of the right adjuvant in order to induce the appropriate type of antigen-specific immune responses in a selective and reliable way. Moreover, the use of adjuvants such the LTK63 and LTR72 mutants, with no or low residual toxicity, holds a lot of promise for the future application of vaccines to the bare skin of humans.