A NOVEL HERPESVIRUS AMPLICON SYSTEM FOR IN-VIVO GENE DELIVERY

For gene therapy approaches to succeed, improved vector systems are ne eded that combine a large carrying capacity with high transduction eff iciency in vivo. Towards this goal we have developed a novel herpes si mplex virus (HSV) amplicon vector, pHE, which contains an HSV-1 replic ation origin (on S) sequence that permit vector replication and packag ing into HSV-1 capsids. The vector also contains the Epstein-Barr viru s (EBV) unique latent replication origin (on P) sequence and a modifie d EBNA-1 gene to allow the vector to be maintained as an episome in tr ansfected E5 helper cells. This system allows for efficient packaging of high-titer vector since the E5 cells are first selected for the pre sence of the pHE vector before helper virus infection. The infectious pHE vector was efficient transgene expression in a variety of human ce ll lines into vitro. Stereotactic injection of pHE vector supernatant into the rat brain resulted in high, localized reporter gene expressio n. Finally, the pHE vector could carry a stable 21 kb DNA payload into HSV virions. This pHE vector system should have a broad range of gene transfer applications.