A problem in the development of vaccines to stimulate the mucosal immune system is to find an appropriate method to deliver the desired antigens to immunoreactive sites such as the small intestine, where the 'gut- associated lymphoid tissue' or 'GALT,' is found. The experimental approach proposed here will utilize poliovirus as a mucosal vaccine because the virus is stable in the intestinal environment, and the attenuated strains are safe for humans. Poliovirus genomes have been constructed which contain the HIV-gag and HIV-pol genes. Transfection of these genomes ('mini-replicons') into cells results in the expression of the HIV-Gag or Pol protein as a fusion protein with the poliovirus capsid proteins. These mini-replicons have been encapsidated by poliovirus capsid proteins provided in trans. Co-infection of these encapsidated mini-replicons with poliovirus type I Sabin (attenuated) or type II Lansing followed by serial passage results in the generation of virus stocks which contain both the encapsidated mini-replicons and the poliovirus type I or type II. The goal of the proposed experiments is to exploit this technology to develop poliovirus as a vaccine for AIDS. The following specific aims are proposed: 1. To express HIV-1 and SIV gp160/env using poliovirus mini-replicons. The full length env gene encoding 856 amino acids will be expressed using a poliovirus mini-replicon. The expressed envelope protein will be characterized by immunological as well as functional assays (capacity to fuse CD4+ cells) to assure that the envelope protein expressed from the mini-replicons corresponds to that expressed from wild type virus. In future studies, we will also express novel env mutants (i.e., secreted envelope, uncleaved precursors) as well as the full length gag gene (Pr55/gag). 2. To test the immunogenicity of encapsidated poliovirus mini-replicons. The appropriate amount of encapsidated mini-replicon and route of immunization (i.e., oral, nasal, intrarectal) to induce a systemic and mucosal immune response will be determined. The studies will use encapsidated mini-replicons in the type II Lansing strain which is adapted for growth in mice. 3. To determine if immunization with encapsidated mini-replicons alone, or in combination with other vaccine formulations, will protect against SIV transmission in a vaginal challenge system. Once we have established the best vaccine strategy using the encapsidated mini-replicons, we will test the vaccine for capacity to protect against vaginal transmission of SIV in collaboration with Dr. John Eldridge and Dr. Preston Marx. The attenuated strains of poliovirus are routinely administered to humans. The incorporation of HIV genes into the poliovirus genome thus establishes a new and novel approach to the development of a vaccine for AIDS.
