This proposal concerns two strategies to prevent herpes keratitis. The first involves blocking the establishment of herpes simplex virus (HSV-1) latency by model anti-HSV vaccines. Vaccinia recombinant viruses will be constructed which express HSV-1 glycoprotein B (gB) under an early vaccinia promoter, gB secreted from the cell membrane, and gC. These vaccinia recombinants will be compared to a previously tested recombinant expressing gB under a late promoter (evaluating protection from HSV-1 corneal disease in experimental animals and the establishment of HSV-1 latency, both in ganglia and in the cornea). It will be determined whether anti-HSV cytotoxic T lymphocytes (CTL) are induced by these vaccines. Limiting dilution analysis and adoptive transfer studies will be done to quantitate and evaluate the protective effect of these CTL's. Peptides corresponding to predicted gB and gC CTL epitopes will be synthesized and tested in vitro and in vivo. The second, more speculative strategy involves the detection during latency of transcripts corresponding to immediate early HSV-1 genes in ganglia and corneal tissue using sensitive and specific polymerase chain reaction (PCR) assays. Sense or anti-sense orientation of transcripts will be determined by the PCR technique. These assays will also be used to detect and quantitate HSV-1 DNA in ganglia and in cornea (of experimental animals and in patients undergoing keratoplasty). A tissue culture neuronal cell model of latency will be used to determine if synthetic oligonucleotides designed to correspond to latency associated transcripts (in a sense or anti-sense orientation) can modulate HSV-1 reactivation in culture. This approach will determine if there is a rational basis for the development of anti-sense therapy for latent HSV-1.
