Abstract

Herpes simplex virus type 1 (HSV-1) amplicons can accommodate foreign DNA of any size up to 150 kbp and, therefore, allow extensive combinations of genetic elements. Genomic sequences as well as cDNA, large transcriptional regulatory sequences for cell type-specific expression, multiple transgenes, and genetic elements from other viruses to create hybrid vectors may be inserted in a modular fashion. Hybrid amplicons use genetic elements from HSV-1 that allow replication and packaging of the vector DNA into HSV-1 virions, and genetic elements from other viruses that either direct integration of transgene sequences into the host genome or allow episomal maintenance of the vector. Thus, the advantages of the HSV-1 amplicon system, including large transgene capacity, broad host range, strong nuclear localization, and availability of helper virus-free packaging systems are retained and combined with those of heterologous viral elements that confer genetic stability to the vector DNA. Adeno-associated virus (AAV) has the unique capability of integrating its genome into a specific site, designated AAVS1, on human chromosome 19. The AAV rep gene and the inverted terminal repeats (ITRs) that flank the AAV genome are sufficient for this process. HSV-1 amplicons have thus been designed that contain the rep gene and a transgene cassette flanked by AAV ITRs. These HSV/AAV hybrid vectors direct site-specific integration of transgene sequences into AAVS1 and support long-term transgene expression.