Expression in plants of the Human Papillomavirus E7 protein and its immunogenicity in mice.

Cutaneous human papillomaviruses (HPV) have been implicated in the etiology of non melanoma skin cancer (NMSC), including basal cell and squamous cell carcinoma (SCC). NMSC incidence has dramatically increased in the last years due to sun exposure and immunosuppression. Although the exact molecular pathways induced following infection by cutaneous HPVs are not defined, their ability to induce malignant lesions in the skin is based on (1) the presence of viral DNA in SCC lesions, (2) the ability of the non structural E6 protein to inhibit UV-induced apoptosis and (3) the induction of tumours in mice models transgenic for the E6-E7 genes (Dong et al., 2005; Schaper et al., 2005). For preventing mucosal HPV infection, prophylactic vaccines based on the viral L1 major capsid protein, assembled into empty virion like particles are already available. Therapeutic vaccines targeting non structural viral proteins, such as the oncogenic proteins E6 or E7 are also under study. However, for cutaneous HPVs, no vaccine strategies have yet been described. In this study, the E7 protein of the cutaneous HPV 8 was modified in its putative pRb binding domain, giving rise to E7-QGD. This protein was transiently expressed in N. benthamiana leaves under the control of the 35S promoter using the agroinfiltration system. For the analysis of transgene accumulation, a polyclonal antibody prepared against a GST-E7 fusion protein produced and purified from E.coli was used. Transgene accumulation was strongly enhanced by the simultaneous agroinfiltration of constructs expressing viral proteins having silencing suppression activity, such as p19 from Carnation Italian ringspot virus and HC-Pro from Potato virus Y. Lyophilised leaves expressing the E7-QGD protein were orally administered to BALB/c mice by gavage. Serum antibody responses were then evaluated by ELISA for the presence of E7-QGD-specific IgGs at 6 and 10 weeks after the first administration. Results of the successful immunisation of mice will be presented and discussed.