This fast track combination Phase I and Phase II application has the overall goal of generating transgenic cattle which express the transgene for a transdominant derivative of a regulatory gene from Bovine Leukemia Virus (BLV). The regulatory gene is from the Rex regulatory region that is required for BLV replication and the generation of new infectious virions. The Rex protein functions to mediate the export and expression of viral RNA's, including singly spliced and doubly spliced RNA's that encode the gag, pol, and env proteins. The use of a transdominant derivative of Rex should function to block the function of the mature, native Rex, and thereby inhibit BLV replication. The basis for this relates to studies with the closely related oncovirus, HTLV-1, and studies with its homologous regulatory Rex gene. Transdominant Rex gene expression in HTLV-1 inhibited HTLV-1 replication in vitro. The goals of the phase I application are to generate high titer, replication incompetent vectors that can deliver the engineered BLV Rex and HTLV-1 Rex to cells, and thereby block replication in vitro. The investigators also propose to engineer into the vectors murine MHC class I molecules that will represent a secondary cell surface antigen with the potential of also generating an immune response in vivo. The justification for this is the transdominant delivered Rex will only inhibit active replication, however, latent provirus will not be affected. The expression of the murine MHC molecules will invoke an immune response in cattle and destroy the latently infected cells. In phase II, the applicants propose to produce transgenic cattle expressing TD Rex and evaluate their ability to prevent progressive infection. Since gestation in cattle is 9 months, the applicants also propose to evaluate whether lymphocytes from transgenic cattle can inhibit BLV infection in vitro. Additional, lymphocytes from naturally infected cattle will also be evaluated to determine if the TD Rex vectors can suppress viral replication in vitro.
