No abstract

Infection with the human papillomaviruses (HPV) types 6 and 11 cause genital warts in man. Current treatments for genital warts are essentially palliative. not treating the underlying infection, and thus recurrences are a common problem. It is thought though that the immune system can control genital warts. Characterisation of the spontaneous regression of genital warts, seen in a proportion of people, shows an increased expression of MHC Class 11 and ICAM-1 on keratinocytes along with a mononuclear cell infitrate dominated by 04' T cells and macrophages. Cantab is developing a therapeutic vaccine designed to stimulate the regression and prevent the recurrence of genital warts. This vaccine is a recombinant fusion protein of the L2 capsid protein to the E7 gene product (L2E7).Believing that a strong CD4' T cell response to HPV6 may eliminate infected cells we have sought to enhance the immunogenicity of our therapeutic vaccine with the use of adjuvants. Monophosphoryl Lipid A (MPL), a detoxified form of lipopolysaccharide, is a promising novel adjuvant that has been shown to stimulate the production of Interferon-y (IFN-y) and enhance T cell responses. We have studied the comparative immunogenicity and reactogenicity of the L2E7 fusion protein in either Alhydrogel, oil in water microemulsions or aqueous adjuvant formulations containing MPL.in two series of experiments in mice. Series 1: Adjuvant formulationsThe immunogenicity of L2E7/MPL formulations was assessed Series 2: Adjuvant formulationsIn all experiments male (B6xCBA) F l mice were used. Every animal received a primary immunisation on day 0 and was boosted 3 weeks later. All immunisations were subcutaneous and injection volumes were 2 x 100p1. Measurements of L2E7 specific immunity were assessed 2 or 3 weeks after the booster immunisation. Serum anti-L2E7 antibody titres were measured by ELISA, in vitro T cell proliferation was measured by incorporation of 3H thymidine and expressed as ACPM, in vitro cytokine production was quantitated by capture ELISA and in vivo DTH reactions to L2E7 were measured as differential increases in ear swelling 24h after intradermal challenee with 10pg L2E7.The addition of MPL to L2E7 adsorbed onto Alhydrogel leads to a 10 fold increase in the in vifro production of IFN-y by antigen specific spleen cells. This response parallels in vitro T cell Proliferation in that it is maximal after immunisation with the middle dose of L2E7 (6pg).The adjuvanting of L2E7 with MPL in a squalene o/w microemulsion primarily enhanced antigen specific serum antibody responses, as shown by increased IgG and IgGlanti-L2E7 antibody titres and the emergence of IgG2a and IgG2bAddition of the Trehalose diester B38 to the d w emulsion broadened the anti-L2E7 antibody response to include IgG3 anti-L2E7 antibodies but did not significantly enhance T cell responses.The simple admixture of L2E7 with aqueous MPL elicited serum anti-L2E7 IgG1, IgG2a & IgG2b antibodies and stimulated IF"-y production but did not stimulate strong T cell proliferative and DTH r...