Design of a Tetracycline Operon Inducible System for the Control of Vaccinia Virus Replication: Implications for Vaccine Development

The use of vaccinia virus (VACV) as a vaccine resulted in the eradication of smallpox in 1979. Characteristics that contribute to the effectiveness of VACV as a vaccine and viral vector include its ability to elicit strong, long-lived humoral and cell-mediated immune responses as a live-replicating virus and to accept large inserts of DNA into its genome. However, adverse events associated with its use as the smallpox vaccine have constrained it from being more widely utilized in vaccines and therapies. We propose to improve the safety of VACV as a live-replicating vector by using elements of the tet operon to control transcription of VACV genes essential for virus growth. This would allow viral replication to be regulated through the addition or removal of tetracyclines. Seven VACV genes were tested in an attempt to control viral replication. For each gene a different recombinant was generated in which the essential gene was placed under the control of tet operon elements. Of the seven VACV genes tested, recombinants utilizing the A6L, A7L, D6R, and F17R genes were successful in regulating viral replication with tetracyclines