Human papillomavirus integration and methylation events and cervical disease progression post-vaccination

• Main Author: Baldwin, Rachel
• Published: Cardiff University 2017
• Subjects: 616.9
• Online Access: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.725707

Human papillomavirus infection is regulated by multiple factors including methylation, viral integration and aberrations in host and viral gene expression. In most patients, the infection is transient and cleared effectively by the host’s immune system but in a minority of cases, the Human papillomavirus infection progresses to neoplasia and cancer if not detected and treated. Assays that detect the presence of Human Papillomavirus infection alone are not good prognostic markers of clinical outcome. Alternative clinical biomarkers that can measure other regulatory factors of Human Papillomavirus infection are required to more accurately predict patient outcome and help direct treatment options specifically to patients at risk of neoplasia and cancer progression. This study aimed to examine several Human Papillomavirus regulatory factors to determine if they would be suitable as clinical biomarkers and explore further the link between molecular changes and associated pathology. This involved development of assays, application to samples obtained from different cohorts of women to ascertain prognostic validity and development of an in vitro system to model the in vivo pathology. Initial work focused on investigating Human Papillomavirus 31, 33, 35 and 51 integration and methylation assays as prognostic biomarkers in young women attending their first routine cervical smear. Results indicated that Human Papillomavirus E2 gene disruption and methylation were not common events and the assays investigated were not suitable biomarkers for predicting clinical outcome in xx young women. The assays were then applied to clinical samples taken from patients with varying grades of cervical disease and high levels of viral methylation were present in women with high grade disease (Cervical Intraepithelial Neoplasia II+) and a correlation between methylation and HPV gene disruption was shown. Novel in vitro organotypic raft cultures of cells from Vulval Intraepithelial Neoplasia and Vaginal Intraepithelial Neoplasia were employed to understand how molecular changes in viral methylation and gene expression correlated with observed pathology. The organotypic raft cultures showed diverse differentiation patterns. No correlation of pathology with viral integration status was detected. However, organotypic raft cultures with high-grade disease morphology all had a higher level of methylation and expression of regulatory genes p16, Ki-67 and Sonic Hedgehog in comparison cultures displaying a histology consistent with low-grade disease. p16 and Ki-67 are already being examined as part of cervical screening triage. The findings presented in this thesis, highlight a need for further research into Human Papillomavirus infection and the molecular changes associated with Sonic Hedgehog gene expression and viral methylation as these show promise as prognostic biomarkers.