A critical challenge for the development of an effective human immunodeficiency virus (HIV) vaccine is the ability of the vaccine to generate a diverse and persistent anti-HIV immune response. Live-attenuated viral variants have been shown to generate such responses, and hence, offer the greatest promise for the development of an HIV vaccine. The effectiveness of live attentuated viral variants could be attributed to their abiltiy to mimic the antigen processing pathways that would occur during infection with wild-type HIV. However, a major concern with the application of live attenuated HiVs as vaccines is their safety. Certain attenuated simian immunodeficiency viruses (SIVS) elicited protective immunity in rhesus macaques after SIV challenge, but subsequently proved to be pathogenic in juvenile monkeys. Thus, there is an urgent need to develop a vaccine which elicits an effective anti-HIV immune response but lacks pathogenic potential. The applicants propose to develop and test in vitro HIV vectors that would replicate with the aid of a helper-expression construct without generating wt-HIV by using ribozymes as a safety mechanism. Such conditionally replicating human immunodeficiency virus (crHIV) vectors would express HIV proteins and, as a result, mimic wt-HIV in the processing of their antigens in vivo. The long-term goal is to determine whether crHIV vectors can be developed to elicit a diverse and persistent anti-HIV immune response without pathogenic consequences. To achieve this goal, the applicants plan to rigorously evaluate the overall feasibility of this approach in vitro by carrying out the following aims: (1) to construct crHIV vector and helper constructs and determine whether the crHIV vector can propagate with the aid of a helper construct in vitro; (2) to assess whether ribozymes contained within crHIV vectors can prevent the emergence of infectious wt-HIV during helper-mediated propagation of crHIV; (3) to determine the degree of antigenic sequence divergence that occurs during propagation of the crHIV vector in vitro; and (4) to determine whether crHIV-containing cells can be infected with wt-HIV and whether crHIV genomes recombine with wt-HIV. The applicants believe these studies will provide the necessary preliminary data to proceed to in vivo testing of this vector system in a non-human primate animal model.
