Results from the RV144 HIV-1 vaccine trial that utilized a systemic heterologous prime-boost immunization regimen consisting of a poxvirus ALVAC gag/pol/gp120 prime combined with a recombinant gp120 boost demonstrated that systemic prime/boost vaccination regimens have the potential to induce modest but significant protection against HIV-1 infection that is presumed to be antibody-mediated. Since HIV-1 infection is predominantly a mucosally-acquired infection, the use of effective mucosal immunization may provide protection superior to that observed in the RV144 trial. The goal of this proposal is to utilize a previously developed systemic prime- boost immunization regimen in non-human primates and develop a new heterologous mucosal prime-boost vaccination strategy to test the hypothesis that optimized mucosal vaccination provides protection against a mucosal SHIV challenge that is superior to protection induced by systemic immunization. To achieve this goal, we will optimize mucosal immunization utilizing three new vaccine components: 1) a novel viral vector, 2) a novel gp120 fusion protein and 3) novel mucosal vaccine adjuvants. The new viral vector that expresses HIV-1 gp140 is a recombinant modified vaccinia Ankara engineered to produce A-type inclusions (MVA/ATI- gp140). The ATI contain mature virus (MV) particles and the use of the MVA ATIs as vaccine vectors is expected to increase mucosal infection with the MVA vaccine vector and allow the recombinant MVA to resist neutralization by MVA-specific antibodies. The novel gp120 fusion protein will contain the adenovirus type 2 fiber (Ad2F) genetically fused to gp120. Previous studies have demonstrated that Ad2F serves as a mucosal targeting ligand to enhance the mucosal immunogenicity of recombinant protein antigens for the induction of neutralizing antibody responses. Additionally, Ad2F may support trimerization of gp120 producing a more native confirmation that may enhance its ability to induce protective antibodies. The novel vaccine adjuvant consist of cationic host defense peptides that have been used to safely induce protective immunity after nasal immunization in non-human primates.