The ultimate goal of this proposal is to produce a vaccine for the mucosotropic virus, canine oral papillomavirus (COPV), which will serve as a model for the development of human papillomavirus (HPV) vaccine. Human papillomaviruses which infect the genital tract mucosa play a critical role in the development of cervical cancer. For example, the majority of human cervical carcinomas (95%) contain and express HPV DNA and it is the expression of two viral oncoproteins, E6 and E7, which appears to be critical for cellular transformation. Despite our detailed knowledge concerning the molecular mechanism of action of these oncoproteins, there is little information available on the biology of papillomavirus infection including the identity viral receptors, the control of viral replication and assembly, and the host immune response to virus and virally-transformed cells. This deficiency derives from a lack of an in-vitro system for propagating the papillomaviruses (which require highly differentiated keratinocytes) as well as the lack of an available animal model to analyze the host factors participating in the response to mucosal infection by papillomavirus. We propose to study the biology of COPV infection in a beagle colony which exhibits a high incidence of oral wart formation as a consequence of viral infection. Our intent is to delineate the role of antibodies in the resistance of animals to this infection which can be produced by the injection of wart extracts (known to contain viral particles). We will sequence the L1 and L2 genes of COPV and clone these genes into expression vectors known (from preliminary results) to produce viral proteins which retain a conformation present in intact viral particles. These viral capsid proteins will be used to screen immune animal sera for the presence of L1 and L2-specific antibodies as well as to induce immunity in susceptible animals. Optimal conditions will be developed for inducing immunity. The ability of L1 and L2 antibodies to inhibit COPV-induced tumors will be evaluated using purified virions derived from wart tissue or, potentially, from viral-producing tumors grown in nude mice. Finally, monoclonal antibodies will be generated against intact virions in order to define the molecular location (L1 or L2, or both) of conformational, neutralizing epitopes on COPV. Due to similarities of COPV and HPV with respect to viral genetic organization, viral structure, capsid protein sequences, and mucosal site of infection, it is very likely that the findings generated with the beagle system will have direct applicability to the development of a human papillomavirus vaccine.
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