Replication-defective bovine adenovirus type 3 as an expression vector

Although recombinant human adenovirus (HAV)-based vectors offer several adv antages for somatic gene therapy and vaccination over other viral vectors, it would be desirable to develop alternative vectors with prolonged express ion and decreased toxicity. Toward this objective, a replication-defective bovine adenovirus type 3 (BAV-3) was developed as an expression vector. Bov ine cell lines designated VIDO R2 (HAV-5 E1A/B-transformed fetal bovine ret ina cell [FBRC] line) and 6.93.9 (Madin-Darby bovine kidney [MDBK] cell lin e expressing El proteins) were developed and found to complement the E1A de letion in BAV-3. Replication-defective BAV-3 with a 1.7-kb deletion removin g most of the E1A and E3 regions was constructed. This virus could be grown in VIDO R2 or 6.93.9 cells but not in FBRC or MDBK cells. The results demo nstrated that the El region of HAV-5 has the capacity to transform bovine r etina cells and that the E1A region of HAV-5 can complement that of BAV-3. A replication-defective BAV-3 vector expressing bovine herpesvirus type 1 g lycoprotein D from the E1A region was made. A similar replication-defective vector expressing the hemagglutinin-esterase gene of bovine coronavirus fr om the E3 region was isolated. Although these viruses grew less efficiently than the replication-competent recombinant BAV-3 (E3 deleted), they are su itable for detailed studies with animals to evaluate the safety, duration o f foreign gene expression, and ability to induce immune responses. In addit ion, a replication-competent recombinant BAV-3 expressing green fluorescent protein was constructed and used to evaluate the host range of BAV-3 under cell culture conditions. The development of bovine E1A-complementing cell lines and the generation of replication-defective BAV3 vectors is a major t echnical advancement for defining the use of BAV-3 as vector for vaccinatio n against diseases of cattle and somatic gene therapy in humans.