This project will test the hypothesis that the oral adjuvant E. coli heat-labile toxin (LT) could be used safely and effectively for enhancing mucosal and systemic immune responses to HIV antigens administered in the rectal and vaginal mucosa of humans. Vaccination strategies that induce durable, high-titered HIV- specific antibodies in rectal and genital tract secretions could prevent sexual transmission of HIV by blocking viral entry at these sites. Current data suggests that local immunization routes would be optimal for generating these antibodies as well as antiviral T helper (TH) cells in the mucosa. However, mucosal adjuvants that augment responses to rectally- or vaginally- administered vaccines have not been identified in humans. Specific AIM I is to confirm uptake and determine the maximal safe dose of LT that can be administered in the rectum and vagina of humans, and to establish an immunogenic dose of HIV p55 gag particles that can be safely co-administered with LT. Groups of women will be evaluated for side effects after small graded doses of LT ranging from 1-5 mug are administered in the rectum or vagina. To confirm LT uptake, local secretions will be screened by ELISA for anti-LT antibodies. Smaller groups of women will then be rectally and vaginally immunized with both maximal safe LT doses and 250 or 500 mug p55 to confirm that the combined preparations are also safe and to establish the rectal and vaginal doses of p55 required for induction of p55-specific antibodies in local secretions. Specific AIM II is to test LT for its ability to enhance systemic and mucosal immune responses and development of immunological memory to p55 co-administered via rectal or vaginal routes. Levels of anti-p55 IgG and IgA antibodies induced in serum and both local and distal secretions of women immunized with p55 alone or p55 with LT will be compared. Induction of TH1/TH2-type responses will be evaluated by quantitating specific IgG subclass antibodies and cytokines secreted in vitro by p55-stimulated peripheral blood mononuclear cells. Memory cell establishment will be assessed by measuring lymphoproliferation in blood using [3H]thymidine uptake assays, and by measuring antibody generated after rectal/vaginal boosting with p55. This study will contribute to knowledge concerning effects of mucosal adjuvants on induction of both local and distal immune responses, design of immunization strategies which produce protective immunity against sexually-transmitted diseases, and mucosal immunogenicity of a promising HIV vaccine subunit component.