AdGFP served as a positive control and showed that 90% of the cells were transduced by this adenovector and expressed the GFP transgene following infection at an MOI of 200 pu/cell. These data support the previous finding that DAF anchor is more efficient to attach antigen to the cell membrane than the native MSP142 anchor in mammalian cells. We next evaluated the immunogenicity of adenovectors expressing the various forms of MSP142 in mice. We observed robust T cell responses 2 weeks after a single immunization for each of the MSP142 expressing adenovectors (Fig. 7a). A second immunization of MSP142 adenovector 6 weeks later did not increase MSP142-specific IFN-γ

responses relative to the 2-week time point (although responses induced by DSA and DS-GM constructs appear MS-275 to be sustained longer in animals that received two doses). The various MSP142 adenovectors differed in their capacity to induce BI6727 MSP142-specific antibody responses in mice. MSP142-specific antibodies were observed in mice after immunization

with a single administration of either AdMSP142-DS or AdMSP142-DSA, but not after one or two doses of AdMSP142-DS-GM or AdMSP142-IC. Interestingly, a second dose of either AdMSP142-DS or AdMSP142-DSA boosted MSP142-specific antibody responses by about 10-fold relative to a single administration of adenovector (Fig. 7b). Adenovector-induced antibody responses were also evaluated in rabbits following two immunizations at an 8-week interval. MSP142-IC was not included in this analysis as it was a poor inducer of antibody responses in the murine studies. The ELISA data with rabbit sera were similar to those from the murine studies. Specifically, the DS and DSA constructs induced the highest responses and

the glycosylation mutant DS-GM induced weak MSP1-specific serum antibody (Fig. 7c). The ability of the MSP142 adenovectors to induce functional antibodies, capable of inhibiting the invasion of erythrocytes by blood stage forms of P. falciparum, was evaluated using GIA. High titers of functional antibodies were induced in rabbits by the adenovectors expressing MSP142. Approximately 60% inhibition was achieved in the standard assay using 2.5 mg/ml of purified IgG from Endonuclease immunized rabbits. The DS and DSA versions of MSP142 induced equally high levels of functional antibodies by GIA ( Fig. 8a) and total antibody by ELISA ( Fig. 8b). We observed similar results using diluted antibody ( Fig. 8c and d). AdMSP142-DS and AdMSP142-DSA performed comparably inducing statistically significant increases in GIA relative to AdNull and AdMSP142-GM (p = 0.0005). There is considerable enthusiasm for the evaluation of adenovirus-based vectors as a gene delivery platform for vaccines. This is driven by findings from different laboratories that adenovectors induce robust and protective T cell responses in multiple animal models of infectious diseases [20], [21], [22], [23] and [24].