Although the relative roles of cell-free and cell-associated HIV-1 in human sexual transmission are unknown, we have found using in vivo SCID mouse and in vitro transwell models of HIV-1 sexual transmission that cell-associated transmission of HIV-1 is highly efficient. These results differ from those reported in the macaque model system, which is likely accounted for by the fact that all of the reported macaque studies make no effort to adjust the environment to the neutral pH at which transmission occurs in the human setting. In our model systems, antibody to the cell adhesion molecule ICAM-1 is remarkably effective in blocking cell-associated transmission. This component of the Program Project proposal will therefore focus on using lactobacillus-based technology to block transmission of cell-associated virus. Preliminary data suggest that the antibody is acting by a mechanism that involves signal transduction rather than by simply blocking the docking function of ICAM-I. We are now hypothesizing that lactobacilli expressing single chain antibody fragments (scFv) can be used as a delivery mechanism for providing a sustained, totally transparent, and, in the case of heterosexual transmission, woman-controlled method for blocking cell-associated virus transmission. To evaluate this hypothesis we have established collaborations with the leading investigators in the world studying the expression of heterologous proteins by lactobacilli. In collaboration with these groups we seek 1) To examine using an in vitro transwell system the differential effects of intact Mab and Fab engagement of ICAM-1 in a cross-linking or a non-cross-linking manner on the movement of HIV-1 infected peripheral blood mononuclear cells across a human epithelial cell barrier. 2) To generate plasmids encoding human ICAM-l-specific scFv followed by their incorpporation into lactobacilli and subsequent testing of the efficacy of the secreted scFv or fixed lactobacilli expressing membrane-bound scFv in the in vitro transwell system. 3) To evaluate the protection efficacy of lactobacilli expressing anti-murine, anti-macaque, or anti-human ICAM-1 scFv in in vivo murine and macaque model systems of transmission of cell-associated and cell-free HIV-1. 4) To develop a chromosomally integrated expression system in Lactobacillus for expression of the selected scFv and subsequent testing in the murine and macaque models.
