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DNA plasmids encoding the open reading frames of canine oral papillomavirus (COPV) nonstructural early genes E1, E2, or E7 protein were delivered into both oral mucosal and cutaneous epithelial sites in beagle dogs using particle-mediated immunotherapeutic delivery (PMID) technology. Control dogs were vaccinated with plasmid encoding either hepatitis B virus surface antigen (HBVs) or COPV L1. Using a prophylactic immunisation protocol, a priming dose of plasmid DNA was followed by a booster dose 6 weeks later. Four weeks after boost, all dogs were challenged with infectious COPV particles. Following viral challenge, as shown previously (M. A. Stanley et al., 2001, Vaccine 19, 2783–2792), mucosal papillomas developed in the negative-control HBVs vaccinated dogs, but all animals in the COPV L1 group were fully protected from disease development. In the early gene-vaccinated groups five of six in the E1-vaccinated dogs, two of six in E2-vaccinated dogs, and three of six in the E7-vaccinated beagles developed oral papillomas. Compared to the HBVs negative-control group the oral papillomas that did develop in the early-gene vaccinated beagles were significantly smaller, shorter in duration, and fewer in number. Taken together the disease burden was markedly reduced and this was statistically significant. In a second experiment one group of animals was vaccinated with plasmid encoding the wild-type COPV E1 gene, and a separate group was vaccinated with plasmid encoding a synthetic codon-optimised COPV E1 gene sequence. None of the codon-optimised E1-vaccinated animals developed papillomas at any challenge site. However, all animals vaccinated with wild-type E1 had papillomas. These data suggest that immunisation by PMID with papillomavirus early genes can significantly impact upon subsequent disease development and that full protection can be achieved using improved vectors encoding codon-optimised gene sequences perhaps emphasizing the importance of antigen load in the generation of protective responses to papillomavirus proteins.

Following challenge with COPV (canine oral papillomavirus), DNA plasmids encoding COPV L1, E1 or E2 protein were delivered into oral mucosal and cutaneous sites in beagles using particle-mediated immunotherapeutic delivery (PMID). Two weeks post-challenge, a priming dose of 8 μg DNA was delivered followed by a booster dose after a further two weeks. A group of control dogs were vaccinated using plasmid DNA encoding Hepatitis B virus surface (HBVs) gene. All of the control animals developed warts at the vast majority of sites (94%). All of the animals given wild type L1, E1, or E2 developed warts at most sites (88%, 75%, and 88%, respectively). The animals given codon optimised E2 however, were protected from wart growth with only one tiny lesion seen on a single animal that persisted for only a few days. The E1 codon optimised group was also significantly protected with a far lower number of smaller warts (48%) that persisted for a shorter duration. These data suggest that therapeutic immunisation by PMID with papillomavirus early genes is effective and emphasizes the importance of antigen load in the generation of protective responses to papillomavirus proteins.

Protection against viral challenge with canine oral papillomavirus (COPV) was achieved by immunisation via particle-mediated DNA delivery (PMDD) of a plasmid encoding the COPV L1 gene to cutaneous and oral mucosal sites in beagle dogs. The initial dose of approximately 9 microg of DNA was followed by two booster doses at 6 week intervals. A similar approach was used to vaccinate a control group of animals with plasmid DNA encoding the Hepatitis B virus S gene. Following challenge at the oral mucosa with COPV all animals vaccinated with the COPV L1 gene were protected against disease. However five of six animals in the control group developed COPV induced papillomas at the oral mucosa. Both cell-mediated lymphoproliferative and humoral antibody responses to the DNA vaccine were observed. Our data indicate that PMDD of plasmid DNA can protect against mucosal challenge with papillomavirus.