Cervical Cancer Cofactors and HPV DNA Integration

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DESCRIPTION (provided by applicant): Human papillomavirus (HPV) infection of the cervix can result in a squamous intraepithelial lesion (SIL) that can progress through increasing less differentiated stages giving rise to cervical cancer (CaCx). The vast majority of CaCx are associated with high risk HPV types. HPV-16 perturbs the function of the tumor suppressor proteins p53 by HPV E6 and pRB by HPV E7 proteins. Abrogation of these activities by HPV-16 oncoproteins can result in the accumulation of mutations and eventually CaCx. Early events in the progression of SIL to CaCx are not well understood. However, it is clear that a change in the physical state of the viral genome (episomal to integrated) likely precipitate CaCx development. Transcripts from integrated virus DNA are more stable than those from episomal virus DNA and the level of viral oncoproteins is higher in cells with integrated sequences. Transcripts of HPV-16 are polycistronic and all early mRNAs contain E7 and E5 sequences. After integration the E5 sequences are lost through fusion of the viral genome with host sequences. We have developed a simple PCR test to distinguish transcripts obtained from integrated or episomal sequences on RNA extracted from residual PreservCyt fluid of the ThlnPrep Pap smear. Only a small percentage of women with SIL will develop CaCx. It has been recognized that cofactors are associated with increased risk for CaCx. These include smoking, methylenetetrahydrofolate reductase (MTHFR) polymorphism, HPV-16 variants, loss of fragile histidine triad (FHIT) gene expression and ChIamydia trachomatis infection. Since integration of the viral genome into the host genome is central to progression to cancer, then women with one or more of these risk factors should have integrated virus DNA in SIL. We have shown that about 22% of women with ASCUS (abnormal squamous cells of undetermined significance) that contain HPV-16 E7 RNA have integrated sequences based on the absence or low levels of E5 transcripts compared to E7 transcripts. We hypothesize that women with integrated HPV-16 sequences have one of more risk factors that are associated with increased risk for cervix cancer. The mechanism of how integration occurs through these cofactors is unknown. We propose to assay for risk factors in women with integrated versus episomal DNA to determine whether the presence of one of more risk factors is associated with integrated sequences. An association will lead to laboratory studies investigating interactions between host and viral genomes giving us a better understanding of events leading to malignant transformation.