A novel approach for herpes simplex virus type 1 amplicon vector production, using the Cre-loxP recombination system to remove helper virus

Helper-dependent HSV vectors (commonly known as HSV amplicons) are able to transfer genes into both dividing and quiescent cells, and thus have the po tential to be widely used as vectors in physiological studies and gene ther apy. Historically, these vectors mere produced by superinfection with a hel per virus that furnished all the trans-acting functions required for amplif ication and packaging of vector genomes into HSV-1 particles. In these syst ems, however, large amounts of potentially harmful helper virus are present in the vector stocks, thus restricting the use of these vectors. New helpe r virus-free packaging systems have been developed that utilize transfectio n of helper functions rather than infection and thus produce safer vector s tocks. The vector titers as well as the amounts of particles obtained with these systems are, however, limited by the impossibility to reamplify the v ector stocks. In this article, we present a novel system for producing larg e amounts of high-titer amplicon vector with low contamination by helper vi ruses. This system is based on the use of the Cre-loxP recombination system , which allows efficient deletion of the packaging signal of an HSV-1 recom binant helper virus (HSV-1-LaL) on Cre-expressing cells (TE-CRE30).