RECOMBINANT HSV VECTORS
Two approaches have been used in the production of Herpes viral vectors (see diagram). In the Amplicon approach, a plasmid containing OriS (the HSV-1 origin of replication), HSV-1 packaging signal and an expression cassette, encoding the therapeutic gene and selection marker under control a viral immediate early promoter, is co-transfected into producer cells with helper virus. The helper virus is a temperature sensitive mutant HSV that encodes all the structural genes. Selection pressure is then applied to the producer cells and viral particles encoding replicon or helper are then produced. The recipient then receives both amplicon and helper particles (see diagram two).
Diagram Two: HSV Amplicons
In the Recombinant approach, HSV is made replication-defective by the deletion of one or more immediate-early genes, e.g. ICP4, which is then provided in trans by a complementing cell line. A gene expression cassette is then introduced into the disabled HSV and used to drive epxression of therapeutic transgene. This second approach is less toxic and can mediate long-term expression of transgene at low concentrations in the brains of mice; however, toxicity in cell culture systems remains a drawback when using recombinant HSV vectors in vitro.
