Immune response to human papillomavirus (HPV) infections is poorly understood because authentic viral proteins are not available for use as antigens. The broad aim of this proposal is to investigate the biology of genital tract HPV-6 and HPV-16 infections, and of HPV-16-associated cervical neoplasia, using type-specific immunological reagents. Unique collections of pedigreed human sera will be utilized in radio- immunoprecipitation assay (RIPA) in order to identify type-specific B cell epitopes on viral proteins synthesized by in vitro transcription and translation (TT-RIPA). The major open reading frames (ORFs) of HPV-6 and HPV-16 will be amplified by PCR (with primers designed to introduce bacteriophage T7 promoter in the amplified sequences) and translated in the rabbit reticulocyte lysate system. Radiolabeled ORF products of HPV-6 and HPV-16 will be reacted in TT-RIPA with serum specimens from appropriate groups (children with recurrent respiratory papillomatosis (RRP) for HPV-6 ORFs; and adults with HPV-16 infections and HPV-16 associated invasive cervical cancer for HPV-16 ORFs) to identify immunoreactive ORFs. Type- specific B cell epitopes on immunoreactive ORFs will be mapped by TT-RIPA of human sera with products of sequential 5'-3' deletions of immunoreactive ORFs, followed by ELISA with synthetic peptides representing minimum immunoreactive regions of the ORF products. Type-specific HPV-6 and HPV-16 reagents will be employed to determine antibody prevalence in selected populations, serologic response to clinical and inapparent infections and correlation between antibody in serum and presence of viral DNA in the genital tract. Serologic markers of cervical neoplasia will be examined for their association with progression or regression of HPV-16 associated cervical cancer and for their prognostic value. T cell lines will be established from peripheral blood mononuclear cells of patients with antibodies to HPV-16 E7 protein and analyzed for the delineation of the T cell epitopes and their fine specificity. Lymphoproliferative response to HPV-16 E7 T cell epitopes will be assessed in women with preinvasive or invasive cervical cancer and related to the stage of the disease and progression or regression of cancer. The proposed investigations will aim to characterize immune responses to HPV infections and to assess their role in the pathogenesis of HPV-associated disease. They may also furnish leads which may be useful in devising strategies for immunologic interventions for prevention and control of HPV-associated cancers.
