Development of a new type of influenza subunit vaccine made by muramyldipeptide-liposome: enhancement of humoral and cellular immune responses

Abstract

The muramyldipeptide (MDP), [6- O-(2-tetradecyl-hexa-decanoyl)-N-acetylmuramyl-l-isoglutamine] can be incorporated into liposomes with haemagglutinin and neuraminidase molecules of influenza virus. Electron microscopic examination revealed that haemagglutinin and neuraminidase subunits were attached to the inner and outer surfaces of lamellar structures of the liposomes, probably through their hydrophobic ends. The addition of cholesterol resulted in much more stable liposomes, which were similar in size and shape to native influenza virus particles. These liposomes enhanced the immunogenicity of haemagglutinin in mice, such that the levels of antibody induced were about 16-fold higher than those of subunit haemagglutinin vaccine alone. Results of proliferation tests with spleen cells from mice and guinea-pigs were consistent with the immunopotentiation of haemagglutinin by liposomes. In addition, the higher antibody levels produced in mice, immunized with the haemagglutinin and MDP-containing liposomes (MDP-virosomes), were maintained for at least 6 months. Enhancement of the cellular immune response, measured by delayed type hypersensitivity reactions, was also observed in the guinea-pigs immunized with MDP-virosome vaccine. Preliminary tests with splenocytes from mice immunized with different vaccines also indicated that the MDP-virosome vaccine induced cytotoxic T-cell activity in these mice. This study revealed that the formation of liposomes with muramyldipeptide enhanced the level and persistence of circulating antibody, and enhanced cellular immunity in guinea-pigs and mice.