Mucosal oncogenic human papillomaviruses (HPV) cause cervical cancer and likely cause a significant fraction of head and neck cancers. We and others have identified HPV-16 isolates that can establish persistent replication and "immortalize" primary human keratinocytes from various sites where HPV associated cancer arise. Assays we developed allow us to evaluate the effect of mutations on individual viral genes throughout the life cycle, including viral gene expression, initial DNA amplification, viral persistence and the influence of epithelial maturation on viral functions using raft cultures. These assays have allowed us to map the structure and regulation of the E1 cistron that is expressed from a novel promoter we have recently defined with a start site at nt 14 (P14). HPV-16 positive carcinomas of the cervix and the head and neck contain and express the viral E6-E7 oncogenic region from a promoter at nt 97 (P97). HPV DNA found in tumors and derived cell lines is frequently disrupted and integrated into the cellular genome and/or contains mutations. These alterations often disrupt the E2 gene, a critical regulator of gene expression. HPV DNAs isolated from carcinomas also frequently contain nucleotide sequence alterations in the viral upstream regulatory region. Our recent studies found these altered HPV genomes could lead to a more aggressive phenotype in our HPV dependent, cancer tissue culture model leading to increases in viral gene expression, virus replication and cell life span and growth rate. The questions this proposal addresses are: 1. What roles do alterations in the upstream regulatory region of HPVs isolated from head and neck cancers have in their pathogenesis? 2. How does E1 expression contribute to establishment of viral persistence? 3. How does E2 expression contribute to establishment of viral persistence?
HPV DNA, present in >90% of cervical cancers and <20% of head and neck carcinomas, is most commonly HPV-16. We propose to study the role of early gene expression and replication in HPV-16. Oral and laryngeal carcinomas are a significant health problem in elderly males. Cervical cancer is a major cause of cancer morbidity and mortality in women. Despite regular cytological screens, many aggressive cases that escape early detection are diagnosed every year. The HPV vaccine will reduce the disease burden of HPV in the future, but most present women veterans will not benefit because they will not have received the vaccine in time. The proposed studies focus on control of HPV-16 replication and gene expression by viral and cellular factors as a critical point in HPV infection and carcinogenesis, and as potential target for future therapeutic intervention.